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Introduction to Genetic Analysis

Introduction to Genetic Analysis

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Een samenvatting van de hoofdstukken die als tentamenstof worden gegeven voor het boek Introduction to Genetic Analysis
Een samenvatting van de hoofdstukken die als tentamenstof worden gegeven voor het boek Introduction to Genetic Analysis

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Introduction to Genetic Analysis



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H1, The genetic revolution in the Life Sciences Genetics is the study of all aspects of genes. Genes are defined as the fundamental units of biological information. Genomics is the study of complete gene sets (called genomes). DNA is biological information encoded as a sequence of nucleotides. DNA is a double helix of two nucleotide chains held together by complementary paring of A-T and G-C. An organism’s complete set of genetic information, encoded in tis DNA, is its genome. The presence of chromosome pairs means that these organisms are diploid, meaning that their nuclei contain two complete copies of the genome and two identical chromosome sets. The number of chromosomes in the basic genomic set is called the haploid number (n). Many eukaryotes such as fungi (schimmels) are haploid. In a diploid, the two members of a chromosome pair are called homologous chromosomes or homologs. The DNA sequences of the members of a homologous pair are virtually the same, even though minor variation in the nucleotide sequence is often present. Page 5 for figure chromosomal landscapes. In diploids, each gene is present as a gene pair. The packing of DNA is achieved by coiling the DNA double helix around molecular spools called nucleosomes (page 7). Each nucleosome is composed of eight proteins called histones. The DNA and associated nucleosomes are together called chromatin, the stuff (grondstof) of chromosomes. One region is very important during cell division, the centromere. The tips of the chromosomes are called telomeres. Inside the mitochondria and chloroplasts are small specialized fraction of eukaryotic genomes found, called the extranuclear genome. The nuclear genomic DNA of eukaryotes is divided into a discrete number of subunits, each coiled around histone proteins in a chromosome. The main functional regions of the DNA are the genes, which are spaced out along the chromosomal DNA. Molecular genetics has shown that biological form is generated by translating the codon sequence of mRNA into the amino acid sequence of protein. The perpetuation (vervolg) of life through time is based on high-fidelity (zeer betrouwbare) replication of a genome’s DNA. A change to the DNA sequence is called a mutation. Certain environmentally induced chemical changes in histones are self-perpetuation, and the altered gene function they cause can also be handed down to descendants. Such nongenetic changes are called epigenetic. Hereditary change is caused mostly by mutations in DNA, but also by epigenetic effects. Natural selection is the process whereby individuals with a particular characteristic (such as better vision) may produce better than others in a given environment. Since these individuals will have more offspring, the relative abundance of individuals with the characteristic in question will increase. Similarity due to the shared ancestry from a common ancestor is called homology. This all-embracing notion of natural selection acting on variation became widely accepted as the theory of evolution. Genetics has made key contributions to understanding evolution, and conversely knowledge of evolutionary homology at the DNA level allow extrapolation from one species’ genetic system to another.

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Y-chromosomal DNA is given trough by the paternal line. Mitochondrial DNA is giving trough by the maternal line. Forward genetics; Mutation  Gene discovery  DNA sequence and function. Reverse genetics; Gene (DNA sequence)  mutation  function. Both forward and reverse genetics work by analyzing mutations and their effects; by showing how a gene goes wrong, we deduce its normal function. DNA cloning means taking a DNA fragment and replicating (amplifying) it many times over until there are many copies so that essentially it can be treated like a reagent in a test tube. One extensively used method for detecting specific macromolecules in a mixture is probing. DNA with Southern blot, mRNA with Northern blot and protein can be detected by Western blot. Finding sets of genes can by using DNA microarrays (page 20). Detecting and amplifying sequences can by using the polymerase chain reaction (PCR). Nucleic acids can be used as labeled probes or primers for detecting homologous nuclei acids on gels, on inorganic surfaces or in solution. Individual proteins can be detected using labeled antibodies. The model organisms are; Escherichia coli (a bacterium), Saccharomyces cerevisiae (baker‟s yeast), Drosophila melanogaster (fruit fly) and the mice. Most genetic studies are performed on one of limited number of model organisms, which have features that make them especially suited for scientific study. Check Summary on page 25. H2. Single-Gene Inheritance Single-gene inheritance patterns may be recognized in the progeny of certain types of controlled matings (paring), which geneticists call crosses. The central components in this type of analysis are mutants, individual organisms having some altered form of a normal property. The normal form of any property of an organism is called the wild type. The use of mutants is sometimes called genetic dissection, because the biological property in question is picked apart to reveal its underlying genetic program, not with scalpel but with mutants. Each mutant potentially identifies a separate gene affecting that property. The genetic approach to understanding a biological property is to discover the genes that control it. One approach to gene discovery is to isolate mutants and check each on for singlegene inheritance patters (specific ratios of normal an mutant expression of the property in descendants). In genetics, the terms character and trait are used more or less synonymously with property. Mendel studied two contrasting phenotypes (verschijnvorm). A phenotype can be defined as a form taken by a character. All the lines used by Mendel were pure lines, meaning that, for the phenotype in question, all offspring produced by matings within the members of that line were identical. How to do crosses? See page 31.

specialized diploid cells called monocytes are set aside to divide to produce sex cells such as sperm and egg in plants and animals or sexual spores in fungi or algae. 5. 2n  n + n + n + n. See page 38 for good summary. heterozygous (Y/y) or homozygous recessive (y/y). In the Y/y plant. and a plant in which the alleles of the pair differ is called a heterozygote. the first cell that develops into a progeny individual. The slash shows that the alleles are a pair. The replication produces pairs of identical sister chromatids. in these organisms. . which become visible at the beginning of mitosis. Hence.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Mendel‟s model for the pea-color example. At the molecular level (right). was as follows.11. Hence. 1. the single DNA molecule of each chromosome replicates. Y/y or y/y). 8. The gene comes in two forms called alleles. Replicated sister chromosomes are together called a dyad. 4. Each plant has a pair of this type of gene. Figure 2. This equal separation has become know as Mendel’s first law or as the law of equal segregation. In genetics generally. each chromosome divides longitudinally into two chromatids (left). 3:1. and. translated into modern terms. Y/y or y/y. An individual can be classified as either homozygous dominant (Y/Y). 6. producing two DNA molecules. gametes fuse randomly. A monohybrid cross is a cross of the type Y/y x Y/y. allelic combinations underlying phenotypes are called genotypes (Y/Y. A fertilized egg. Sometimes a heterozygote for one gene is called a monohybrid. and the two nuclear divisions that accompany them are called meiosis. on for each chromatid. four cells are produced. Because there are two divisions. Most eukaryotes have a sexual cycle. the members of a gene pair separate equally into the eggs and sperm. and the results are haploid. A plant can be either Y/Y. At fertilization (bevruchting).com . If the gene is phonetically called a “wye” gene. Mendel’s inheritance is shown by any segment of DNA on a chromosome: by genes and their alleles and by molecular markers not necessarily associated with any biological function. then the two alleles can be represented by Y and y. All 1:1. the Y allele dominates and so the phenotype will be Y.Stuvia. the phenotype of the Y/y plant defines the Y allele as dominant and the y allele as recessive. A plant with a pair of identical alleles is called a homozygote. See page 42 for a good overview of differences between nuclear division in haploid and diploid cells and between mitoses and meiosis. 7. In meiosis. The unit comprising the pair of synapsed dyads is called a bivalent. 3. Meiosis only takes place in diploid cells. The four chromatids that make up a bivalent are called a tetrad. See page 39 for information about single-gene inheritance in haploids. regardless of which of the alleles they bear. A hereditary factor called a gene is necessary for producing pea color. Two sequential cell divisions take place. and 1:2:1 ratios are diagnostic of singlegene inheritance and are based on equal segregation in a heterozygote. 2. a single gamete contains only one member of the gene pair. is called a zygote.

in a heterozygote (+/M). Choose biological property of interest Find mutants affecting that property Check mutants for single-gene inheritance Identify time and place of action of genes Zero in on molecular nature of gene by genomic (DNA) analysis The cross of an individual of unknown heterozygosity (for one gene or more) with a fully recessive parent is called a testcross. Dominance was defined earlier as the phenotype shown by a heterozygote. Gene discovery by single-gene inheritance is sometimes called forward genetics. geneticists more often apply the term to alleles. when the mutant first shows up in the population. a 1:1 ratio will result. Other genes are haploinsufficient. In a heterozygote (+/m. a null mutant allele will be dominant because. In haploids assigning dominance is usually not possible. Because this number indicates single-gene inheritance. In general. then another approach is needed). the single wild-type allele cannot provide enough product for normal function. and the recessive individual is called a tester. they are sometimes called leaky mutations. it works in the following sequence: 1. all progeny will show the dominant phenotype. In such cases. we can conclude that the mutant is cause by a recessive alteration (verandering) in a single gene. 5. it will be in the heterozygous state. The outcome of F1 is all the same en the outcome of F2 is an ratio of 3:1. Other mutant alleles reduce the level of enzyme function. If the individual is homozygous. but it is like Mendel‟s experiment. 2. Most mutations alter the amino acid sequence of the gene’s protein product. If the individual is heterozygous. resulting in reduced or absent function. 4. for F2 you have more options. To discover genes there is a standard procedure to cross the mutant with wild type (if the mutant is sterile (onvruchtbaar). One copy of a haplosufficient gene provides enough gene product to carry out the normal transactions of the cell. .Stuvia. in practice. because some wild-type function seems to “leak” into the mutant phenotype. m is a null) the remaining copy encoded by the + allele provides enough protein product for normal function. de ander is geïnactiveerd door een mutatie).com . See page 47. only a single „dose” of mutant allele is necessary. The principles of inheritance (such as the law of equal segregation) can be applied in two directions: (1) inferring genotypes from phenotypic ratios and (2) predicting phenotypic ratios from parents of know genotypes. formally it is the phenotype that is dominant or recessive. so in most cases. For a dominant mutation to be expressed. The F1 has an ratio of 1:1. 3.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Many of mutant alleles are of a type generally called null alleles: the proteins encoded by them completely lack function. See page 48. but. Mutations elsewhere in the gene may have no effect on enzyme function and are called silent mutations. Recessiveness is observed in mutaties in genes that are functionally haplosufficient (als een diploïd organisme maar een functioneel kopie van een gen heeft. Hence.

and so they called pseudo autosomal regions 1 and 2. who bear the recessive allele masked in the heterozygous condition. A member of a family who first comes to the attention of a geneticist is called the propositus. . as well as different ratios in reciprocal crosses. 2. They are both carrying A/a.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal In most of the cases. a polymorphism is the coexistence of two or more common phenotypes of a character. consisting of one X and one Y. In human. the two X chromosome pair segregate like autosomes. they are autosomal-like. an autosomal (page 56) recessive disorder is generally revealed by the appearance of the disorder in the male and female progeny of unaffected parents. Many more males than females show the rare phenotype. This because a female can only inherit the genotype if both the parents bear the allele (example. Sexlinked inheritance regularly shows different phenotypic ratios in the two sexes of progeny. Let‟s look at a pedigree for an interesting human case. 3. If the recessive allele is very rare. one at each end. and so each egg receives one X chromosomes (homagemetic sex). See page 51 for X and Y chromosomes. See page 53. In the sense that these regions are homologues. In general. XAXa * XaY) whereas a male can inherit the phenotype when only the mother carries the allele (XAXa * XAY). such pedigrees typically show the following features: 1. Males have nonidentical pair. The human X and Y chromosomes have two short homologous regions. Mutant alleles in the differential region of the X chromosome show a single-gene inheritance pattern called X linkage. the morphs are common. A dimorphism is the simplest type of polymorphism. females have a pair of identical sex chromosomes called the X chromosomes.com . nor will they pass the condition to their descendants. genes in the differential regions are said to show inheritance patterns called sex linkage. In males half the sperm contain X and half Y (hetrogametic sex). with just two morphs. Populations of plants and animals (including human) are highly polymorphic. At meiosis in females. This because only the Y is given trough. X linked recessive disorder. None of the offspring of an effected male show the phenotype. Mutant alleles of the few genes in the differential region of the Y chromosome show Y linkage. sex is determined by special pair of sex chromosomes. almost all persons showing the phenotype are male. None of the sons of an affected male show the phenotype under study. they also show affected men and women transmitting the condition to equal proportions to their sons and daughters. The alternative phenotypes of a polymorphism (morphs) are often inherited as alleles of a single autosomal gene in the standard Mendelian manner. by definition.Stuvia. but all his daughters are “carriers”. The other 22*2=44 chromosomes are called autosomal chromosomes. See page 59 for example. In natural populations of organisms. In human pedigrees. The interpretation of pedigrees for polymorphisms is somewhat different from that of rare disorders because. Pedigrees of mendelian autosomal dominant disorders show affected males and females in each generation. The Y chromosome is considerably shorter than te X. so progeny with the recessive a/a will be produced. Contrasting morphs are often inherited as alleles of a single gene.

A dihybrid is a double heterozygote such as A/a * B/b. Affected heterozygous females married to unaffected males pass the condition to half their sons and daughters. The product rule states that the probability of independent event both occurring together is the product of their individual probabilities. See page 93 for an example of autosomal gene independent assortment in combination with X-linked genes. See page 87 for table. The general superiority of multiple heterozygotes is called hybrid vigor. E is the expected number in a class. Ratios of 1:1:1:1 and 9:3:3:1 are diagnostic of independent assortment in one and two dihybrid monocytes respectively. . The sum rule states that the probability of either of two mutually exclusive events occurring is the sum of their individual probabilities. gene assortment when the hybrid undergoes meiosis breaks up the favorable allelic combination. Mendel’s second law the principle of independent assortment) states that gene pairs on different chromosome pairs assort independently at meiosis. the proportion of heterozygotes is reduced to (1/2)8 = 1/256. r/y. From studying dihybrid crosses ((A/a * B/b) * (A/a * B/b)) Mendel came up with his second important principle of heredity. For calculating risks in pedigree analysis see page 63. and thus few members of the next generation have it.com . In the last fase of meiosis the chromosome pair act independently. They all show in the F1 the same character and in F2 a 9:3:3:1 ratio. See page 79 for information about dihybrid crosses. such pedigrees show the following features: 1. a process that can be used to create pure lines for research or other applications. Affected males pass the condition to all their daughters but to none of their sons. and look at the summary on page 64. r/r*y/y) and the outcome was an 1:1:1:1 (see page 91 for a figure about independent assortment of chromosomes at meiosis explains Mendel‟s ratio) ratio of R/Y. Mendel testcrossed the F1 dihybrid (R/r*Y/y) with a tester of genotype (recessive. H3. R/y. The independent assortment of genes at meiosis is one of the main ways by which an organism produces new combinations of alleles. r/Y.Stuvia. Repeated selfing leads to an increased proportion of homozygotes. see page 88. and the alleles of two heterozygous gene pairs are said to show independent assortment. However. 2. The production of new allele combinations is formally called recombination. O is observed number in a class. Inheritance patterns with an unequal representation of phenotypes in males and females can locate the genes concerned to one of the sex chromosomes. The random fusion of these gamets results in the 9:3:3:1 ratio. Independent assortment of genes. X2 calculation by Sum( O – E )2 / E for all classes.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal X linked dominant disorder. After 8 generations of selfing.

Any product of this process is called a recombinant. In contrast nuclear genes separate the cytoplasm equally. which are haploid meiotic products with new combinations of the alleles carried by the haploid genotypes that united to form the meiocyte.Stuvia. Meiosis generates recombinants. but we can infer these genotypes by using the appropriate techniques. Variation and assortment of polygenes can contribute to continuous variation in a population.com . we use pure-breeding diploid parents because they can produce only one gametic type. To know the input gametes. . Organelle populations that contain mixtures of two genetically distinct chromosomes often show segregation of the two types into the daughter cell at cell division. Organelle genes show their own special mode of inheritance called uniparental inheritance: progeny inherit organelle genes exclusively form one parent but not the other. A recombinant frequency of 50% indicates that the genes are independently assorting and are most likely on different chromosomes. This process is called cytoplasmic segregation. In most cases that parent is the mother. We cannot detect the genotypes of input or output gametes directly. a pattern called maternal inheritance.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Meiotic recombination is any meiotic process that generates a haploid product with new combinations of the alleles carried by the haploid genotypes that united to form the meiocyte. To detect recombinant output gametes. Why the mother? Because the organelle chromosomes are located in the cytoplasm and the male and female gametes do not contribute cytoplasm equally to the zygote (the egg takes almost everything). Mutant female * wild-type male  progeny all mutant Mutant male * wild-type female  progeny all wild-type Variant phenotypes caused by mutations in cytoplasmic organelle DNA are generally inherited maternally and independent of the Mendelian patterns shown by nuclear genes. Examples of relative gene size and spacing in mitochondrial DNA (mtDNA) and chloroplast DNA (cpDNA) are shown on page 100. See page 95 for a good figure on this subject. The interacting genes underlying hereditary continuous variation are called polygenes or QLT‟s. Other message see page 103 See page 105 for the summary. we testcross the diploid individual and observe its progeny.

By determining the frequency of recombinants. they do not assort independently but produce recombinant frequency of less than 50 percent. Recombination maps of chromosomes are usually assembled two or three genes at a time. generally. Dus linkage analyses is dat als de plaatsen van de twee of meerdere genen op hetzelfde chromosoom zitten. Dus 50%= 2e wet van Mendel. the greater the proportion of recombinants that would be produced. with the use of a method called linkage analysis. arising from two crossovers. Double-recombinant classes can be used to deduce the extent of this interference. When two genes are close together on the same chromosome pair (that is that they are linked). A crossover is the breakage of two DNA molecules at the same position and their rejoining in two reciprocal recombinant combinations. One chiasmate is equal to 50 cM. map the loci of genes that have been identified by mutant phenotype showing single-gene inheritance. Mapping Eukaryote Chromosomes by Recombination. onafhankelijke rangeren van genen. they are assembled by quite different procedures yet are used in a complementary way. Hence. See page 122. crossovers inhibit each other somewhat in an interaction called interference. Recombination-based maps. In fact. the alleles on any one homolog are physically joined (linked) by the DNA between them. they mean that the loci of those genes are on the same chromosome. This because we treated the two rarest classes of progeny with respect to the recombination of v and cv.u. We discuss physical maps in chapter 15. not chromosomes.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal H4. a recombinant frequency of less that 50 percent is a diagnostic for linkage. Chiasmata are the visible manifestation of crossovers. Crossing over is between chromatids.com . ends up higher than expected. The two smallest classes are the DCO. recombinants are produced by crossovers. we can obtain a measure of the map distance between the genes. are the crossovers in adjacent chromosome regions independent events or does a crossover in one region affect the likelihood of there being a crossover in an adjacent region? The answer is that.The total m. For a good example of a three-point testcross see page 126. hence. . Now we have the map. Double crossovers can include three or four chromatids. and. I = 1 – ( observed frequency or number of double recombinants / expected frequency or number of double recombinants) The last u calculate by the two chances * number of total population. When geneticists say that two genes are linked. Bij minder dan 50% is het aanneembaar dat ze gelinkt zijn en dus niet onafhankelijk rangeren. en dus de allelen op een homoloog fysiek contact kunnen maken met het DNA tussen hen. Dus hoe groter de afstand tussen twee genen (op het zelfde chromosoom) hoe groter het recombinatie percentage is. For linked genes. which are the topic of this chapter. Sturtevant defined one genetic map unit (m.u. The greater the distance between the linked genes. conversely.) as the distance between genes for which is 1% = 1 cM. we can see that these two rare classes are in fact double recombinants. The basic types of chromosome maps are currently used in genetics.Stuvia. The two classes with the most progeny are the parental types. We can ask.

Another important type of dominant mutation is called a dominant negative. Harmful mutations of haplosufficient genes are often dominant. Mutations in genes that encode units in homo. the single wild-type allele cannot provide enough product for normal function. So it is an dominant mutation. For most genes. In such cases. as in a heterozygote. a single copy is adequate for full expression (haplosufficient). A fully dominant allele will be expressed when only one copy is present. and their null mutations are fully recessive. In this situation the homozygous dominant cannot be distinguished from the heterozygote (example = PKU). The genes encoding the enzymes of a specific pathway constitute a functionally interacting subset of the genome. To see if a gene is essential. the expression of both alleles of a heterozygote. de ander is geïnactiveerd door een mutatie). See page 202. An allele that is capable of causing the death of an organism is called a lethal allele. The genetic approach that reveals the interacting genes for a particular biological property is briefly as follows. whereas the alternative allele will be fully recessive. The simplest type of dominance is full. In cases in which the mutants infer. such as pigment. But we can al . acting through “spoilers” (verknoeide) proteins. 1. m is a null) the remaining copy encoded by the + allele provides enough protein product for normal function. If mutants alleles from different genes interact. Recessiveness is observed in mutaties in genes that are functionally haplosufficient (als een diploïd organisme maar een functioneel kopie van een gen heeft. Another variation on the theme of dominance is codominance. A total of six different genotypes can be made (see page 203 and also for anemia). which contain three alleles. dominance. One copy of a haplosufficient gene provides enough gene product to carry out the normal transactions of the cell. Collect single mutants and test for dominance. The occurrence of the intermediate phenotype suggests an incomplete dominance. in a heterozygote (+/M).com . a null allele is tested for lethality.or heterodimers can behave as dominant negative. each wild-type allele generally produces a set dose of its protein product and the number of doses corresponds with the concentration of a chemical made by a protein. a null mutant allele will be dominant because. then we infer that the wild-type genes interact normally as well. A clear example is seen in the human ABO blood groups. or complete. It can be explained by in incomplete dominance.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal H6. a modified 9:3:3:1 Mendelian ratio will often result. Other genes are haploinsufficient.Stuvia. In a heterozygote (+/m. Gene interaction The known mutant alleles of a gene and its wild-type allele are referred to as multiple alleles or an allelic series. Chemical synthesis in cells is by pathways of sequential steps catalyzed by enzymes.

Epistasis is inferred when a mutant allele of one gene masks the expression of a mutant allele of another gene and expresses its own phenotype instead. So penetrance is defined as the percentage of individuals with a given allele who exhibit the phenotype associated with that allele. Reasons not to: the influence of the environment. they measure. terwijl 12:3:1 dominante epistatias impliceert. If they have the wild-type phenotype than they are on different alleles (called complemented). When two independently derived recessive mutant alleles producing similar recessive phenotypes fail to complement. The next step is to observe whether the progeny have the wild-type phenotype. absence of either gene function leads to absence of the end product of the pathway. that is. See page 210 and PP. Los van dosis dus. The terms penetrance and expressivity quantify the modification of gene expression by varying environment and genetic background.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal 2. Hierdoor kun je dus verhoudingen als 7:9 krijgen enzovoorts. A 9:7 ratio suggests interacting genes in the same pathway. Construct double mutants to see if genes interact (see page 212). resulting in normal wild-type phenotype. or other interacting genes of the sublety of the mutant phenotype. expressivity measures the intensity of the phenotype. the percentage of cases in which the gene is expressed and the level of expression. A range of modified 9:3:3:! F2 (from selfing of a dihybride F1) ratios can reveal specific types of gene interaction. Expressivity measures the degree to which a given allele is expressed at the phenotypic level. Een 9:3:4 verhouding impliceert recessief epistatias. Mutant alleles called suppressors cancel the expression of a mutant allele of another gene. then the recessive mutations must be alleles of the same gene. Complementation is the production of a wild-type phenotype when two haploid genomes bearing different recessive mutations are united in the same cell. respectively. In a diploid. The same or at a different locus? A quick approach often used is the complementation test. they must be alleles of the same gene.com .Stuvia. the complementation test is performed by intercrossing two individuals that are homozygous for different recessive mutations. . When the progeny are not wild type. 3.

producing diploid gametes. Polyploids are often larger an have larger component parts than their diploid relatives. 5n = pentaploid. All chromosomes comparing to each other occur in the same ratio. page 241. resulting in a condition called aneuploidy. Allopolyploids form only between closely related species and the chromosome sets are homeologous (partly homologous). Large-Scale chromosomal changes Chromosome mutations can be detected by microscopy. and see this page also for a calculation about genetic ratios. monoploid zygotes fail to develop. 4n = tetraploid. In most species. The deleterious recessive alleles are masked by wild-type alleles in the diploid organism but are automatically expressed in a monoploid derived from a diploid.If chromosomes in a tetraploid pairs as bivalents or quadrivalents. Zaadloze bananen. Polyploids with odd numbers of chromosome sets. An individual of a normally diploid species that has only one chromosome set is called a monoploid (all insects and schimmels). and changes in parts of chromosome sets. homozygous diploids. the chromosomes segregate normally. Autopolyploids have multiple chromosome sets originating from within one species. This chapter is all going about eukaryotes. such as triploids. together called a “genetic load”. resulting in a condition called aberrant (afwijkende) euploidy. Polyploids Polyploids are individual organisms that have more than two chromosome sets. Monopolids are sterile (geen nageslacht). or by combination of all techniques. . or at can be spontaneously. An amphidiploid (dubbele diploid).com . 6n = hexaploid. Allopolyploids have sets from two or more different species. Colchicine may be applied to generate a tetraploid from a diploid. F1 is sterile because the chromosomes of the two different gametes are not as homologous. see page 239.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal H7. Genetics can create new plant lines by producing monoploids with favorable genotypes and then doubling their chromosomes to form fertile. 3n = triploid. Euploid means that you have the basic sets of chromosome number. The reason is that virtually all members of a diploid species carry a number of deleterious recessive mutations. by genetic or molecular analysis. etc. Allopolyploid plants can be synthesized by crossing related species and doubling the chromosomes of the hybrid or by fusing diploid cells. See page 240. are sterile or highly infertile because their gametes and offspring are aneuploid. Organisms that have more or fewer than the normal number (normal = haploid or diploid) of sets are aberrant euploids. Changes in chromosome number are of two basic types: changes in whole chromosome sets. We can use colchicine to create a diploid.Stuvia.

Why are aneuploid so much more abnormal than Polyploids? Why does aneuploidy for each chromosome have its own characteristic phenotypic effects? Why are monosomics typically more severely affected than are the corresponding trisomics? The answers seem certain to be a matter of gene balance.Stuvia. A human chromosome complement of 44 autosomes plus a single X produces a condition known as Turner syndrome. The relationship between the number of copies of a gene and the amount of the gene‟s product made is called a gene-dosage effect. The orientation of a segment within the chromosome can be reversed (inversion) or it can be moved to a different chromosome (translocation). Both breakage and rejoining and crossing over between repetitive DNA can lead to deletion etc. For a good summary on deletion and everything see page 253. With a multigenic deletion several too many genes are missing. the ratio can be different (in an trisomic 2:3. turner. first division nondisjunction occurs. An aneuploid is an individual organism whose chromosome number differs from the wild type by part of chromosome set. balanced if the chromosome gene change order but do not remove or duplicate any DNA (inversion or a reciprocal translocation) and unbalanced change the gene dosage of a chromosome segment (a deletion or duplication). The lethality of large heterozygous deletions can be explained by gene imbalance and the . If crossing over fails for some reason. A small deletion within a gene is called an intragenic deletion and has the same effect as that of an null mutation. Represented as XO. There are two types of rearrangement. Geeft geen problemen voor nageslacht en er wordt altijd maar 1 X of Y afgeschreven. see page 250-251).De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Aneuploids Aneuploidy is the second major category of chromosomal aberrations in which the chromosome number is abnormal. sterile) or trisomic (2n + 1. The only chromosomal rearrangements that survive meiosis are those that produce DNA molecules that have one centromere and two telomeres. A chromosome can be lost (deletion) or doubled (duplication). The combination XXY results in Klinefelter syndrome. Persons with this syndrome are males who have lanky builds and a mildly impaired IQ and are sterile.com . Changes in chromosome structure are called rearrangements. down. In an aneuploid. Down syndrome caused by the appearance of an extra chromosome 21. Aneuploid organism result mainly from nondisjunction in a parental meiosis. Monosomic (2n-1. Aneuploidy is nearly always deleterious because of gene imbalance: the ratio of genes is different from that in euploids. Another important cause of rearrangements is crossing over between repetitive (duplicated) DNA segments called nonallelic homologous recombination (NAHR). In an euploid cell the gene balance is 1:1. With XXX and XYY most males and female are fertile. fertile). and this difference interferes with the normal function of the genome. Crossovers are needed to keep bivalents paired until anaphase 1. Page 246.

reduced recombinant frequency. to form a reciprocal translocation. See page 256. See page 257 for information about de cri du chat syndrome. In produces progeny carrying an almost complete extra copy of chromosome 21. one chromosome twists once at the ends of the inversion to pair with the its untwisted homolog. they do not change the overall amount of genetic material.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal expression of deleterious recessives. a segment of a chromosome is cut out. Recall that. Because recessive alleles seem to be showing dominance in such cases.Stuvia. Inversion spanning the centromere are called pericentric. Two central questions.com . Becuause inversions are balanced rearrangements. is a genetic diagnostic clue to the presence of a translocation. In meiosis. it is called paracentric. Variegation (spreading) can result from a gene‟s unusual proximity to heterochromatin in a translocation heterozygote. A product that lacks one centromere is called a acentric chromosome. To see adjacent and alternate segregation and their products see page 264. Is there a genetic (in)balans? What happens in meiosis? Summary 269. The apparent (schijnbare) linkage of genes normally known to be on separate nonhomologous chromosomes. to create an inversion. in this way. The duplicate regions can be located adjacent to each other (tandem duplication) or the extra copy can be located elsewhere in the genome (insertional duplication). individuals with inversions are generally normal if there are no breaks within genes. and so they do not result in gene imbalance. the translocation responsible is of a type called a Robertsonian translocation. See figure on page 267 for cancer by somatic translocation (due the gene is now on a different promoter). We consider only reciprocal translocations. If the centromere is outside the inversion. and a likely explanation is an inversion spanning most of the dp – cn region (see page 263). The failure of a small deletion on the normal homolog to pair creates a visible deletion loop. Inversion heterozygote is when inside a diploid cell that contain one normal chromosome set plus on set carrying the inversion. flipped an reinserted. forming an anaphase bridge. the paired homologs form a visible inversion loop. Heterozygous reciprocal translocations are diagnosed genetically by semisterility and by the apparent linkage of genes whose normal loci are on separate chromosomes. In down syndrome. and reduced fertility because of unbalanced or deleted meiotic products. psuedolinkage. the simplest type. The main diagnostic features of heterozygous inversions are inversion loops. See figure 7. PMS and the Williams syndrome. the effect is called pseudodominance. We have seen that. two chromosomes trade acentric fragments created by two simultaneous chromosome breaks. A product with two centromeres is called a dicentric and it will simultaneously pulled to opposite poles at anaphase. Another clue to the presence of a deletion is that the deletion of a segment on one homolog sometimes unmasks recessive alleles present on the other homolog. which is illustrated on page 265. .24 for information about segmental duplications. Something is reducing crossing over in a region.

siRNA‟s restrain (bedwingen) transposable elements in plants. and piRNA‟s perform the same function in animals. if any. DNA contain purine A and G. C---G). but unlike DNA – can catalyze biological reactions (ribozymes). Classes of functional RNA. while both siRNA‟s and piRNA‟s prevent the spread of transposable elements to other chromosomal loci. Small interfering RNA’s (siRNA’s) and piwi-interacting RNA’s (piRNA’s) help protect the integrity of plant and animal genomes. RNA‟s can be grouped into two general classes. r-RNA molecules are the major components of ribosomes. of most is currently unknown. Bovenstaande worden door weinig genen geproduceerd maar wel in grote aantallen waardoor een groot % van het RNA in de cel uit deze RNA-typen bestaat. which are large macromolecular machines that guide the assembly of the amino acid chain by the mRNA‟s and tRNA‟s. RNA has ribose sugar in its nucleotides. MicroRNA’s (miRNA’s) have recently been recognized by scientists to have a widespread role in regulating the amount of protein produced by many eukaryotic genes. not a double helix like DNA. RNA: Transcription and Processing The transaction of DNA and RNA take place through the matching of complementary bases and the binding of various proteins to specific sites on the DNA or RNA. RNA – like protein. Bij processing.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal H9.Stuvia. Long noncoding RNA’s (Inc/nc RNA’s) were recently found to be transcribed from most regions of the genomes of humans and other animals and plants. Some snRNA‟s unite with several protein subunits to form the ribonucleoprotein processing complex (spliceosome). - - . siRNA‟s inhibit the production of viruses. C---G) and RNA contain purine A and G en pyrimidine C and U (A--U. Differences between RNA and DNA: RNA is usually a single-stranded nucleotide chain. While a few IncRNA‟s play a role in classic genetic phenomena such as dosage compensation (H13). en pirymidines C and T (A--T. t-RNA molecules are responsible for bringing the correct amino acid to the mRNA in the process of translation. the function. We refer to the other class as functional RNA because the RNA does not encode information to make protein.com . rather than the deoxyribose (H ipv OH op 2nd C atom) found in DNA. Small nuclear RNA’s (snRNA’s) are part of system that further processes RNA transcripts in eukaryotic cells. One class of RNA encodes the information necessary to make polypeptide chains (proteins) called mRNA. Genes that are important for regulation of RNA and protein levels in the cell.

Transcription is asymmetrical: only one strand of the DNA of a gene is used as a template for transcription.3’. In some way the same can be said for transcription. . those that encode proteins (mRNA) and those that are functional as RNA (ncRNA’s). called a transcription bubble (luchtbel). in the direction opposite the direction of transcription. RNA polymerase usually binds to a specific DNA sequence called a promoter. This multisubunit complex is composed of the five subunits of the core enzyme plus a subunit called the sigma factor. The replication of DNA in eukaryotes. RNA processing (snRNA). located close to the start of the transcribed region. DNA replication is more complex in eukaryotes in large part because there is a lot more DNA to copy. The promoter is referred to as upstream of the initiation site because it is located ahead of the initiation site. The template strand is the strand which is used by RNA polymerase 11. the nontemplate strand is also called the coding strand. The DNA is said to be transcribed into RNA. where transcription begins. Termination The transcription of an individual gene continues beyond the protein-ending segment of the gene. rRNA). Functional RNA’s participate in a variety of cellular processes including protein synthesis (tRNA.Stuvia. although more complicated. the regulation of gene expression (miRNA) and genome defense (si and pi RNA). is very similar to the replication of DNA in prokaryotes. A downstream site would be located later in the direction of transcription. There are three different stages of transcription Initiation In prokaryotes. This strand is on the 3’. TATAAT (-10) and TTGACAT (-35).com . creating a 3’UTR at the end of the transcript. because eukaryotes retain many of the events associated with initiation. it maintains a region of single-stranded DNA. The intervening part is referred to as the 5’untranslated region (5’UTR). The bacterial RNA polymerase that scans the DNA for a promoter sequence is called the RNA polymerase haloenzyme. and the RNA is called a transcript.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal There are two general classes of RNA’s. The protein-encoding part of the gene usually begins at an ATG sequence. and termination in prokaryotes. and RNA is synthesized in to the 5’. Elongation proceeds until RNA polymerase recognizes special nucleotide sequences that act as a signal for chain termination. but the initiation site. elongation. Elongation In a way. is usually well upstream of this sequence.5’ orientation.

See chapter 13. In this way. 3. RNA’s capable of catalysis are called ribozymes. The function of GTF’s is to attract the core RNA polymerase 11 so that it is positioned to begin RNA synthesis at the transcription start site. (2) splicing to eliminate introns and (3) the addition of a 3‟polyA tail. Processing takes place during RNA synthesis. Alignment of the snRNP’s result from hydrogen bonding of their snRNA molecules to the complementary sequences of the intron. See page 324. whereas it is virtually naked in prokaryotes. After transcription has been initiated. The template for transcription. Eukaryotic pre-mRNA is extensively processed before being transported as mRNA to the cytoplasm for translation into protein. Before the RNA leaves the nucleus. it must be modified in several ways. called the carboxyl tail domain (CTD) and this is strategically located near the site at which nascent RNA will emerge from polymerase. One gene can encode more than on polypeptide when its pre-mRNA is alternatively spliced. The spliceosome is composed of several snRNP‟s that attach sequentially to the RNA. TATA box is about -30 bp upstream. The machine which is responsible for all these processing is called the RNA splicing machinery. This processing includes (1) the addition of a cap at the 5‟end. Here is binding of the TATA-binding protein (TBP). Eukaryotic promoters are first recognized by general transcription factors. Intron removal and exon joining are catalyzed by RNA molecules. Some introns are selfsplicing. this is why multiple RNA polymerase 11 enzymes can simultaneously synthesize transcripts from a single gene. is organized into chromatin in eukaryotes. RNA polymerase 11 transcribes all protein-encoding genes and some snRNAs. introns are removed and exons spliced together. . RNA polymerase 11 dissociates form most of the GTF‟s.com . In eukaryotes. 1. A 5’cap and 3’polyA tail are added. the reactants are properly aligned and the two splicing reactions can take place. taking up positions roughly as shown on page 327. Before processing the mRNA is called the pre-mRNA. the intron itself catalyzes its own removal.Stuvia. The initiation phase ends and the elongation phase starts after the CTD has been phosphorylated by one of the GTF‟s. genomic DNA. Also eukaryotic depending on general transcription factors (GTF’s). also called RNA processing. 2. in these cases.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Transcription is more complicated in eukaryotes for three primary reasons. The larger eukaryotic genomes have many more genes to be recognized and transcribed. the snRNA´s of the spliceosome catalyze the removal of introns from pre-mRNA. The GTF‟s and the RNA polymerase 11 core constitute the preinitiation complex (PIC). One of the subunits of RNA polymerase 11 contains a protein tail.

recognizes ds RNA molecules and cleaves them into 22/23 nucleotide products. They may also recruit RNA polymerase 11 more directly through binding. RISC binds a short RNA and unwinds it to form biologically active siRNA. See page 329 for a good figure. an siRNA silences the gene that produces it. called siRNA. In bacteria. The biologically active single strand miRNA binds to RISC and guides it to complementary sequences in protein-coding mRNA’s. The second set of regulatory proteins consists of transcription factors that bind to cis-acting regulatory sequences in the DNA called enhancers (versterkers). RNA polymerase 11 complex and general transcription factors (GTF‟s). still bound to RISC. where RISC either represses translation or promotes mRNA degradation. which is degraded. To initiate transcription. the packaging of DNA with nucleosomes prevents transcription unless other regulatory proteins are present. however. In eukaryotes. A second machine. See page 441.com . The miRNA. promoters and promoter-proximal elements are bound by transcription factors that affect the expression of many genes. has a very different origin and function from miRNA‟s. See page 332 for a good figure. binds to a short ds RNA and unwinds it into the biologically active single-stranded miRNA. H13. One machine. The siRNA targets RISC to a perfectly complementary mRNA. thus silencing the expression of the foreign DNA. RISC then represses the translation of these mRNA‟s into protein or removes the polyA tail. Dicer detects double stranded RNA that forms between antisense and sense RNA an processes it into short RNA’s. Regulation of Gene expression in Eukaryotes. an enhancer will act in only one or a few cell types in a multicellular eukaryote. which binds to doublestranded RNA’s. which hastens mRNA degradation. A second type of short RNA. RNA polymerase can usually begin transcription unless a repressor protein blocks it. called Dicer. In contrast to miRNA‟s. interacts with DNA sequences called promoter-proximal elements near the promoter of a gene. Often. The binding of sequence-specific DNA-binding proteins to regions outside the promoters of target genes is a common feature of eukaryotic transcriptional regulation. Antisense RNA is frequently formed in response to the insertion of foreign DNA into the genome. called RISC (RNA-induced silencing complex).Stuvia.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal miRNA‟s halt translation from targeted genes by two biological machines. UAS= upstream activator sequence. Enhancers are bound by transcription factors that control the regulation of small subsets of genes. . Generally speaking. miRNA’s are processed from longer RNA pol 11 transcripts by Dicer. binds to complementary mRNA‟s. These regulatory proteins expose promoter sequences by altering (veranderen) nucleosome density (dichtheid) or position.

It can be modified by acetylating or methyling. Eukaryotic activators recruit RNA polymerase 11 to gene promoters through two major mechanisms. These protruding ends are called histone tails. transcription factor 11D. as well as other eukaryotic regulators. The changing of nucleosome position is referred to as chromatin remodeling. Chromatin remodeling changes nucleosome density or position and is an integral part of eukaryotic gene regulation. a large multiprotein complex that. One subcomplex. Chromatin can be dynamic. so the presence or absence of these groups can carry a tremendous amount of information. There are 44 lysine residues available to accept acetyl groups. activators can recruit proteins that having modify chromatin structure allowing RNA polymerase 11 and other proteins access to the DNA. having separable domains for DNA binding. First activators can interact with subunits of the protein complexes having roles in transcription initiation. upstream activation sequence) meestal stroomopwaarts van de transcriptie start positie. The mediator complex is an example of a coactivator. it recruits the TF11D complex and. the covalent modification of histone tails is said to be a histone code. Een activatie domein en een bind domein. RNA polymerase 11 to the promoter. directly interacts with RNA polymerase 11 to recruit it to gene promoters. The histone proteins are organized into the core octamer with their amino-terminal ends making electrostatic contacts with the phosphate backbone of the surrounding DNA.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Cel specifieke transcriptie wordt bereikt door de activiteit van transcriptie activators en repressor. Second. The binding to TBP in a side of the activation domain it can activate gene expression.Stuvia. histones are almost identical in all eukaryotic organisms. in turn. Eukaryotic transcriptional activators often work by recruiting parts of the transcriptional machinery to gene promoters. nucleosomes are not necessarily in fixed positions on the chromosome. in turn. whereas inactive genes are underacetylated. die binden aan specifieke DNA sequenties (UAS. Many eukaryotic transcriptional regulatory proteins are modular (bestaande uit verschillende modules) proteins. Through this binding interaction. .com . The ability of Gal4. In yeast and in multicellular eukaryotes. cell-type-specific patterns of gene expression are governed by combinations of interacting transcription factors. A second way a gene can be activated is by interacting with the mediator complex. Histones are known to be the most conserved proteins in nature. binds to the TATA box of eukaryotic promoters through the TBP. that is. The activity of eukaryotic transcriptional regulatory proteins is often controlled by interactions with other proteins. to function in a variety of eukaryotes indicates that eukaryotes generally have the transcriptional regulatory machinery and mechanisms in common. Evidence has been accumulating for years that the histones associated with the nucleosomes of active genes are rich in acetyl groups. Er zijn twee domeinen actief verantwoordelijk voor activatie eiwit. For this reason. See page 446. activation or repression and interaction with other proteins.

the enhancer-blocking insulators prevent the enhancer form activating transcription at that promoter. Enhancers contain binding sites for many transcription factors. Enhancers are bound by transcription factors that control the regulation of small subsets of genes. These interactions result in a variety of response. So the histone code is genetic inherent. When positioned between the enhancer and the promoter. Eukaryotic enhancers can act at great distances to modulate the activity of the transcriptional apparatus. Active genes that are relocated to genomic neighborhoods that are heterochromatic may be silenced in the heterochromatin spreads to the genes. A regulatory element. ze trekken beide eiwitten aan om translatie te starten. histone acetylation and deacetylation promote and repress gene transcription. which bind and interact cooperatively. . See page 455. These activities are recruited to genes by sequence-specific activators and repressors.com . but it also prevents the binding of other proteins.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal In most cases examined. respectively. To prevent such promiscuous activation. De chromatine status bepaald dus of een gen wel of niet geactiveerd wordt. Generally speaking. Activator is dicht bij de promoter. promoters and promoter-proximal elements are bound by transcription factors that affect the expression of many genes. namelijk chromatine structuur en het methylatiepatroon DNA. terwijl een enhancer ver van de promotor kan liggen. Een van de allelen blijft voor altijd uit geschakeld om over expressie te voorkomen. In replication. The imprinting control region (ICR) is unmethylated in female gametes and can bind a protein forming an insulator that blocks enhancer activation. Enhanceosomes help recruit the transcriptional machinery for a short-term activation of genes in a chromatine environment. See page 456 and 457. regulatory elements called enhancer-blocking insulators have evolved. De methyleringsgraad van lysine in histonen speelt een grote rol in de vorming van hetero chromatine. Enhancer-blocking insulators are a fundamental component of a phenomenon called genomic imprinting. where they direct the coding of adjacent newly assembled histones to form complete nucleosomes. old histones with their histone codes are distributed randomly to the daughter strands. So C methylation would be associated with inactive regions of the genome. Highly condensed heterochromatic regions have fewer genes and low recombination frequencies than do the less-condensed euchromatic regions. Methylation of the ICR in male germ cells prevents CTCF binding. such as an enhancer.Stuvia. They also recruit chromatin remodelers. that can act over tens of thousands of base pairs could interfere with the regulation of nearby genes. Hoe toegankelijk het DNA is voor tarnscriptie is zelf ook epi-genetisch bepaald door de chromatine status. including the recruitment of additional co-activators and the remodeling of chromatin. Most of the unmethylated dinucleotides are found in clusters near gene promoters. Er worden twee dingen overgeërfd. The chromatin of eukaryotes is not uniform.

Zie pagina 468. Om het RNA te vertalen is een veel grotere machine nodig dan bij DNA replicatie en transcriptie. tRNA moleculen zijn moleculaire adapters. aminoacyl-tRNA synthetases. ze linken aminozuren aan de codons. both alleles of a gene are expressed independently. called fibrous proteins. een RNA stuk van 16kb is hiervoor verantwoordelijk. Dat komt omdat deze complementaire tRNA moleculen moet vangen. they are called globular proteins. Specifieke nucleotiden in zowel het anticodon als de aminozuur accepterende arm zorgen ervoor dat het correct tRNA gesynthetiseerd wordt door het correct enzym en dus de correct aminozuurgroep eraan komt te zitten.com . Herkenning en het toevoegen van aan aminozuur aan de juiste tRNA wordt gedaan door enzymen. De kleine subunit linkt (decoding center) de tRNA‟s aan de codons op het mRNA en de grote subunit katalyseert (peptidyltransferase center) de formatie van een peptidebinding die aminozuren aan elkaar binden.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal See page 466 and 467 for an example of differentially imprinting in males and females. Proteins and their synthesis. meaning that each of the 64 triplets must have some meaning within the code. For most diploid organisms. In these cases. Many proteins are compact structures. See message page 345 and 350. For the code to be degenerate some of the amino acids must be specified by at least two or more different triplets (wobble). Enzymes and antibodies are among the best-known globular proteins. epigenetic mechanisms silence a single chromosomal locus or ony copy of an entire chromosome. The genetic code is degenerate. Ook zijn er meerdere tRNA voor de aminozuren (meerdere codons coderen voor een aminozuur). 348) en aan de ene kant hebben ze een aminozuur bij zich. Ze zijn 80 nucleotiden lang. hair. Deze machines zijn ribosomen. are important components of such structures as skin. Ze komen elkaar dus tegen op het mRNA en het mRNA wordt als het ware door het ribosoom heengetrokken.Stuvia. Eukaryotische en prokaryotische ribosomen zijn qua structuur en vorm hetzelfde. Sommige tRNA‟s kunnen zijn zo gemaakt dat alleen een binding met de eerste 2 posities van het codon genoeg is. vasthouden en de aminozuren die de tRNA moleculen met zich mee brengen aan elkaar moet rijgen. . In de meeste organismen is er voor elk aminozuur 1 verschillinde synthetases. Een van de twee x-choromosomen moet worden stilgelegd om overdosis te voorkomen. aan de andere kant een anticodon. H10. Ze hebben beide een grote en een kleine subunit. Proteins with linear shape. and tendons. Genomic impringint and X inactivation are examples of only a single allele being available for expression. A peptide bond is formed by the linkage of the amino end of one amino acid with the carboxyl end of another amino acid. Het anticodon bind aan het complementaire codon in het mRNA. gevormd als een klaverblad (pag. en er dus een mismatch wordt toegestaan op de 3e positie.

Ribosomen zijn gemaakt van 2/3e rRNA en 1/3e eiwit. De beginstap is zeer belangrijk om in het goede „reading‟ frame te komen. Niet alleen de bindingsplaatsen bij de kleine subunit zijn gemaakt door rRNA‟s.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Elk ribosoom bevat een bindingsplaats voor mRNA en drie bindingsplaatsen voor tRNA (EP-A). een aminozuur wordt toegevoegd aan de eiwitketen. het laatste punt is waarbij een cel kan beslissen of het mRNA wel vertaald moet worden. Dat heeft weer tot gevolg dat de eiwitketen loslaat en hierna laten de ribosomen het mRNA los. Daarna schuift het ribosoom iets op en het gebruikte tRNA molecuul zit nu in de E-site waar het wordt uitgeworpen. Het tRNA molecuul komt binnen bij de A-site door baseparing aan te gaan met het complementaire mRNA codon. In bacteriën is dit anders omdat deze geen 5‟cap hebben om de kleine subunit te vertellen waar het moet beginnen met het zoeken naar een startcodon. Deze wordt vaak verwijderd. In eukaryoten kan alleen het begintRNA sterkt binden met de P-site van de kleine subunit. Translation is carried out by ribosomes moving along mRNA in the 5’-3’direction. molecuul. Het aminozuur wordt dan gelinkt aan de eiwitketen die wordt vastgehouden door de vorige tRNA (op de P-site). Dit is essentieel omdat mRNA‟s van prokaryoten vaak voor meerdere eiwitten coderen. In zowel eukaryoten als prokaryoten wordt het einde van een eiwit zichtbaar gemaakt door een stopcodon. Een prokaryotisch ribosoom kan gelijk binden aan een startcodon op het mRNA als het maar voorafgegaan wordt door specifieke nucleotidensequentie.v.p. organ. . ook het katalyserende gebied in de grote subunit (23S RNA). An incoming amino acid becomes bonded to the amino end of the growing polypeptide chain in the ribosome. Pag. Ook is het belangrijk omdat het. Er zijn hiervoor geen tRNA‟s die binden maar eiwitten die ontsnappings factoren genoemd worden binden aan elk stopcodon dat de A-site bereikt  dit leidt ertoe dat een watermolecuul i. The proteome is enriched by two cellular processes: the alternative splicing of premRNA and the posttranslational modification of proteins.Stuvia. Translatie begint met het codon AUG en een speciaal tRNA (het begintRNA. Belangrijk is om te zien dat de grote subunit zich eerst verplaatst en daarna pas de kleine subunit. Deze „geladen’ subunit bindt aan het 5‟ eind (door 5‟cap) van een mRNA molecuul en verplaatst zich net zolang in de richting van het 3‟eind (poly-A staart) totdat het een AUG tegenkomt. Dan verdwijnen alle translatie begin factoren. 352. The proteome is defined here as the complete set of proteins in an organism. and their anticodons bind to mRNA codons exposed on the ribosome. die vanaf het begin aan de kleine subunit zaten om zo ruimte te maken voor de grote subunit.com . tissue or cell. Een ribozym is een RNA molecuul dat een katalytische rol speelt in een biochemische reactie. en is anders dan het normale tRNA molecuul dat methionine meebrengt). Alle eiwitten hebben dus allemaal methionine aan het N-eind (het eind dat als eerst wordt gesynthetiseerd). A set of tRNA molecules bring amino acids to the ribosome.

New synthesized proteins contain a signal sequence at its amino-terminal end.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Alternative splicing can lead to the synthesis of multiple proteins (called isoforms) with different combinations of functional domains. Most cutting is done with the use of bacterial restriction enzymes. thus promoting protein activation or degradation. You would expect that the folding is going wrong because the aqueous environment inside the cell does not favor the correct folding of most proteins. Another key property of som restriction enzymes is that they make “sticky ends”. A restriction enzyme will cut the DNA into a set of restriction fragments determined by the locations of the restriction sites. The long-size chromosome DNA molecules of genomic DNA must be cut into fragments of much smaller size before they can be inserted into a vector. modify amino acid side groups.Stuvia. Some posstranslational events. their addition to a protein usually changes protein conformation.com . tissue or cell. . You can do this in vivo or in vitro. and most often it is an entire genome. Replications of DNA segments is called amplification. Other posstranslational mechanisms recognize amino acid signatures in a protein sequence and target those proteins to places where their activity is required inside or outside the cell. including enzyme activity. The two most common. It can be a DNA palindrome. More than 300 modifications are possible to amino acid side chains after translation. The interactome is the name given for whole sets of protein-protein interactions in an organism. Most eukaryotic proteins are inactive unless modified after translation. The in-vitro approach is called PCR and both ways leads to recombinant DNA due DNA cloning. Fragmenst of the donor DNA are isnterted itno a specially designed plasmid or bacterial virus that will “carry” and amplify the gene of interest and are hence called vectors. Single-strand paring in this way also called hybridization. The sample with the gene of interest is called the donor DNA. Phosphorylation Because phosphate groups are negatively charged. The most important posttranslational event is the folding of the nascent protein into its correct three-dimensional shape. protein-protein interactions and protein-DNA interactions. Uniquitin The modification targeting a protein for degradation is the addition of chains of multiple copies of a protein called uniquitin. The addition and removal of phosphate groups serves as a reversible switch to control a variety of cellular events. See page 371. H11. The proteins are folded correctly with the help of chaperones. Gene isolation and Manipulation. such as phosphorylation or ubiquitination.

mRNA is often a preferable straing point in the isolation of a gene. Enzymatic conversion of mRNA into (reverse transcriptase) cDNA allows for the isolation of a gene copy without introns. Page 382-383. There are two types of probes. The most popular vector for cloning very large DNA inserts is the bacterial artificial chromosome BAC (100-200kb).The task of isolating a clone of a specific gene begins with making a library of genomic DNA or cDNA. Donor and vector DNAs that are the products of digestion by the same restriction enzyme that produces sticky ends can be joined efficiently and ligated. Page 380.if possible. See page 375. The polylinker has several alternative restriction sites into which donor DNA can be inserted.Stuvia. As a first step. See page 379.with this library of DNA inserts. (1) those that recognize a specific nuclei acid sequence (autoradiogram) and (2) those that recognize a specific protein (antibody‟s). followed by the amplification ot these molecules as either plasmid chromosomes or phages. Identify clones from the library that produce transformed cells with the dominant a+ phenotype. Insertion of DNA into pUC18 is detected by inactivation of the B-galactosidase function of lacZ resulting in an inability to convert the artividial substrate X-Gal into a blue dye. Alternatively.com . Bij een PCR van cDNA zijn veel fragmenten blunt. Transform cells of recessive-mutant-cell-line a. The plasmid vector PUC18 has been designed for use as a vector for DNA cloning. donor DNA that is the product of PCR or cDNA synthesis requires the addition of sticky ends prior to insertion into a vector. Standart vectors can insert about 25 to 30 kb. Bij het inbrengen van een gen in een recombinante DNA plasmide is het veel handiger om sticky ends creërende restrictie enzymen te gebruiken. and many of them generate single/stranded sticky ends suitable for making recombinant DNA.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Genomic DNA can be used directly for cloning genes. it has an promotor and an drug-resistant gene. Fosmids are vectors that can carry 35. Vandaar dat sticky ends handiger zijn. omdat deze allemaal kunnen ligeren met een vector is dit een heel inefficiënt proces. The general outline of the procedure of functional complementation or mutant resque. One alternative method is to creat PCR products with sticky ends by using specially designed PCR primers that contain restriction endonuclease recognition sequences at their 5‟end. Recover the a+ gene from the successful bacterial or phage clone. The polymerase chain reaction uses specially designed primers for direct isolation and amplification of specific regions of DNA in a test tube. restriction enzymes cut DNA into fragments of manageable size. Make a library contain wild-type a+ recombinant-donor DNA inserts.to 45 kb inserts. Gene cloning is carried out through the introduction of single recombinant vectors into recipient bacterial cells. enriched for sequences containing the gene in question. See page 377. .

A random set of clones from this region was placed into the correct order. This was done in part by using a technique called a chromosome walk (page 392). a natural plasmid from a soil bacteriu. Beofre complete genome sequences were available. The fragments in distinct size classes wil form district bands on the gel. RNA molecules can also be detected in a mixture on a gel. investigators can use fine mapping to narrow the search for the gene of interest. and its strategy is to use the genetic position to isolate the gene underlying the trait. or even bacteria. The bacterium causes what is known as crown gall disease. DNA can now be interoduced from other species of plants. This process is know as positional cloning. Thansgenesis can introduce new or modified genetic material into eukaryotic cells. No longer is genetic diversity achieved solely by selecting variants within a given species. with the use of a technique called Southern blotting (DNA). A probe can identify one fragment in this mixture. The most extensively used method for detecting a molecule within a mixture is blotting. or mRNA’s for measurement of the size specific DNA or RNA. A cloned DNA segment can be sequenced by characterizing the end bases of a serial set of truncated synthetic DNA fragments. molecular cloning of genes for genetic disorders such as systic fibrosis or certain cancers was a prodigious undertaking. The bands can be visualized by straining the DNA with ethidium bromide. in which the infected plant produces uncontrolled growth called tumors or galls. which causes the DNA to fluoresce in ultraviolet light. See page 393 for fine mapping. the isolation of defective diseasecausing genes begins with the genetic mapping of the disease trait. this by Nothern blotting. With tightly linked markers flanking the trait in hand. which starts with gel electrophoresis to separate the molecules in the mixture. each terminated at different positions corresponding to the incorporation of a dideoxynucleotide. Recombinant-DNA techniques that depend on complementarity to a cloned DNA probe include blotting and hybridization systems for the identification of specific clones. The sequencing technique to sequence whole genomes is called dideoxy sequencing or Sanger sequencing.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal A cloned gene can be selected form a library by using probes for the gene’s DNA sequence or for the gene’s protein product or by complementing a mutant phenotype. producing genetically modified organisms (GMOs). The basic idea is to use the sequence of the nearby landmark as a probe to identify a second set of clones that overlaps the marker clone containing the landmark but extends out from it one of two directions. restriction fragments. A vector routinely used to produce transgenic plants is derived from the Ti plasmid. Even with access to the sequences of entire genomes.Stuvia.com . . animals.

1. Millions of individual DNA fragments are isolated and sequenced in parallel during each machine run. One major challenge facing a genome project is sequence assembly. each employ three strategies that have dramatically increased throughput (doorstroom). building up all of the individual reads into a consensus sequence. 2. this is. Characterizing whole genomes is fundamental to understanding the entire body of genetic information underlying the physiology and development of living organisms and the discovery of new genes such as those having roles in human genetic diseases.com . Genomic sequences are obtained. The sequences of overlapping areas are assembled into units called sequence contigs (sequences that are contiguous. 4. Genomes en genomics. and software make it possible to detect the products of sequencing reactions in extremely small reaction volumes. In one application. A gene may be inactivated by substituting an inactive gene for the normal gene.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Gene targeting enables researchers to eliminate a gene or modify the function it encodes. or touching). a sequence for which there is consensus (agreement) that it is an authentic representation of the sequence for each of the DNA molecules in that genome.Stuvia. However different systems have been devolved which differ in certain details. overlapping small segments. DNA molecules are prepared for sequencing in cell-free reactions. Read the sequence of each small segment. . called gene replacement. a mutant allele can be repaired by substituting a wild-type allele for the mutant one in its normal chromosomal location. cameras. without cloning in microbial hosts. such a targeted inactivation is called a gene knockout. 2. Germ-line transgenic techniques have been developed for all well-studied eukaryotic species. The current general strategy for obtaining and assembling the sequence of a genome is called whole-genome shotgun sequence (WGS). Break the DNA molecules of a genome up into thousands to millions of more or less random. Computationally find the overlap among the small segments where their sequences are identical. 3. Advanced fluid-handling technologies. We seen in chapter 11 how this works (traditional WGS). H15. The goal of next-generation WGS is the same as that of traditional WGS-to obtain a large number over overlapping sequence reads that can be assembled into contigs. Continue overlapping ever larger pieces until all the small segments are linked. 3. These techniques depend on an understanding of the reproductive biology of the recipient species. 1. the approach is to.

De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal One of the most widely used approach. The DNA molecules in the template library are amplified into many copies. Predictions of mRNA and polypeptide structure from genomic DNA sequence depend on the integration of information from cDNA sequence.C. 5. So called processed pseudogenes are DNA sequences that have been reversetranscribed from RNA and randomly inserted into the genome. After each synthesis a laser is applied resulting in the removal of the 3‟blocking group and the probe. Species evolve and traits change through changes in DNA sequence.T. In addition to full-length cDNA sequences. and codon bias. 4. each fluorescently labeled with a different color and attached with a blocking group. The landscape of eukaryotic chromosomes includes a variety of repetitive DNA segments. polypeptide similarities. Ninety percent or so of human pseudogenes appear to be of this type. The sequencing of each bead is performed using a novel „sequencing-by-synthesis‟ chemistry termed pyrosequencing. The second step is the identification of the most closely related genes. The four bases then compete for binding sites on the template DNA to be sequenced and non-incorporated molecules are washed away. but by using the PCR. These segments are difficult to align as sequence reads. These short cDNA sequence reads are called expressed sequence tags (EST‟s). The first step in comparing genomes is to decide which species genomes to compare. in elk welletje zitten meerdere dezelfde strengen. called homologs. There are more than 19. DNA molecules are first attached to primers on a slide and amplified so that local colonies are formed.com . there are large data sets of cDNA‟s for which only the 5‟ or the 3‟ends or both have been sequenced. which are ORF‟s or partial ORF‟s that may at first appear to be genes but are either nonfunctional or inactive due to the manner of their origin or to mutations. 2. dA/C/G/TTp‟s worden na elkaar toegevoegd en zichtbaar gemaakt door sulfurylase en luciferase. . In addition to the identification of conserved regions. such as homopolymers and repetitive sequences.G) of reversible terminate bases are added. 1. The four types of (A. 3. binding-site predictions. A detectable fluorescent color specific to one of the four bases is then visible. not by growing colonies as for traditional genomic libraries. have three stages. Illumina sequencing can be used to sequence difficult regions. Komt er op neer. The evolutionary history of a group is called a phylogeny. developed by 454 Life Sciences Corporation. 3.000 pseudogenes. A DNA template library of single-stranded DNA molecules is constructed. It works as follows. 1. allowing for sequence identification and the beginning of the next cycle.Stuvia. comparative genomics has the potential to reveal how species diverge. 2. See page 516 and 517 for more information. but also for whole-genome and region sequencing.

2. One of the greatest surprises that has emerged form comparing individual human genomes is the degree to which humans differ not merely at one base in a thousand. Determining which genomic elements have been gained or lost during evolution requires knowledge of the phylogeny of the species being compared. They arose when genes within a genome were duplicated. Genes that are related by gene-duplication events in a genome are called paralogs.com . Checken met bank . met reverse-transcriptase omgezet naar cDNA en er wordt een fluorescent (rood. weefsel met tumorcellen) label aan gemaakt. entire genes or sets of genes. Rood: Is de mate van expressie hoger in de tumorcellen. Dit zijn stukjes van 25 nucleotiden en er zijn zoveel spots dat wiskundig gezien alle mogelijke DNA-sequenties wel aan een probe hechten. However. Alles wordt weggewassen en de volgende mogelijkheden ontstaan. Zie pagina 535. More than 90 percent of the mouse and human genomes can be portioned into corresponding regions of conserved synteny. and interaction of gene products is termed functional genetics. Deze twee gelabelde cDNA producten worden samengevoegd en op de plaat gegoten waarna het cDNA zal hybridiseren aan de genen in de spots die een complementaire sequentie hebben. The presence or absence of genes ofrten correlates with organism lifestyles. Een DNA-microarray is een chip met daarop grote hoeveelheid spots met in elke spot een andere “probe”. The mouse and human genomes contain similar sets of genes. Geneticists have been studying the expression and interactions of gene products for the past several decades. many homologous genes belong to families that have expanded (and contracted) in number in the course of evolution. bijv. 4. controlegroep). often arranged in similar order. 3. Fylogenie (van het Griekse φυλη (phulè = 'volksstam') en γενεσις (genesis = 'wording')) is de studie van de ontstaansgeschiedenis van een groep organismen. These homologous genes are at different genetic loci in the same organism. The global approach to the study of function. 1. but in the number of copies of parts of individual genes. Deze wordt veelal grafisch weergegeven in een fylogenetische stamboom. De fylogenie onderzoekt niet zozeer de overeenkomsten tussen verschillende organismes. Een ander monster wordt met een andere kleur gelabeld (groen. Geel: Is de mate van expressie vergelijkbaar. kan dit betekenen dat de tumor afhankelijk is van die veranderde expressie. These copy number variations (CNVs) include repeats and duplications that increase copy number and deletions that reduce copy number. Het mRNA wordt geëxtraheerd. Een fylogenie is de beschrijving van hoe de ene groep organismen is ontstaan uit andere groepen. expression. where the order of genes within variously sized blocks is the same as their order in the most recent common ancestor of the two species. These genes would have been inherited from a common ancestor and are referred to as orthologs. Groen: Is de mate van expressie van dit gen hoger is in de gezonde cellen. Als een gen vaak een veranderede expressie heeft bij dat kankertype.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Some homologs are genes at the same genetic locus in different species.Stuvia. maar de evolutie van die organismen uit gemeenschappelijke voorouders.

This mechanism is called RNA interference (RNAi). to what yeast sequence. Each domain is connected to a different protein. 2. 1. 2. The intact Gal4 protein is a transcriptional activator which has two separate functions ( a binding domain and an activation domain). If the two proteins interact.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal One of the most common ways of studying protein-protein interactome uses an engineered system in yeast cells called the two-hybrid test. 4. if so. Starting from the available gene sequences is referred to as reverse genetics. yo use a antibody that reacts specifically with the encoded protein. Studying the protein-DNA interactome using chromatin immunopreciptitation assay (ChIP). The Gal4 gene is divided between two plasmids so that one plasmid contains the part encoding the DNA-binding domain and the other plasmid contains the part encoding the activation domain.Stuvia. Page 537. Targeted mutagenesis is the most precise means of obtaining mutations in a specific gene and can now be practiced in a variety of model systems. The two hybrid plasmids are then introduced into the same yeast cell. a gene for one protein under investigation is spliced nest to the DNA-binding domain (bait). and on a other plasmid a gene for another protein is spliced nest to the activation domain (target). including mice and flies. One of the most exciting discoveries of the past decade or so has been the discovery of a widespread mechanism whose natural function seems to be to protect a cell form foreign DNA. 3. which detects physical interactions between two proteins. Suppose that you are investigating whether tow proteins interact (page 536). . The DNA bound can be analyzed directly or be amplified into many copies by PCR to prepare for DNA sequencing. 1. they will join the two domains together. The activator will become active and start transcription of the reporter gene. The strategy of the two-hybrid system is to separate the two domains of the activator encoded by Gal4 making activation of a reporter gene impossible. 4. but almost the same a gene knock-out. On one plasmid. 3. You want to know whether this protein binds to DNA and. RNAi-based methods provide general ways of experimentally interfering with the function of a specific gene without changing its DNA sequence. To separate the fragment containing your protein-DNA complex from other. Page 539. You have isolated a gene form yeast an suspect that it encodes a protein that binds to DNA.com . You can treat yeast cells with a chemical that will cross-link proteins to the DNA. 5. two antibiotic resistant. Thus the two domains must be in close proximity in order for transcriptional activation to take place. Break the chromatin into small species.

com .Stuvia.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal .

Stuvia. en soms zelfs weer in klassen opdelen. . For some traits. Continuous traits typically have complex inheritance involving multiple genes plus environmental factors. this is the part of the phenotypic variance that is due to genetic differences among individuals in a population within a single generation. Traits such as height that show a continuous range of variation and do not behave in a simple Mendelian fashion are known as quantitative or complex trait.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal H19.o.v.v. Fenotypische variatie = Genetische variatie + mulieuvariatie. the individuals in a population can be sorted into discrete groups or categories. Such traits are known as categorical traits. H2 = broad-sense heritability. The term complex trait is often preferred to continuous or quantitative trait because it includes all the types of traits with which quantitative genetics is concerned. Mathematically. variance and normal distribution. H2 is not useful for interpreting differences in trait means among populations. including the mean. Fenotype = genotype + milieu. dan kan zelfs een monogene eigenschap kwantitatief overerven! Indien de milieuvariantie relatief klein is t. H2 provides a measure of the extent to which differences among individuals within a population are due to genetic versus environmental factors. The fields of quantitative genetics studies the inheritance of complex traits using some basic statistical tools. Vx = Vg + Ve Indien de bijdrage van milieuvariantie relatief groot is t. Estimates of H2 apply only to the population and environment in which they were made. A complex trait can be either a discontinuous trait such as the presence or absence of a disease condition or a continuously variable trait such as height in humans. These traits often exhibit simple inheritance. H2 = Vg / Vx (Vg+Ve) Broad-sense heritability H2 is the ratio of the genetic (Vg) to the phenotypic (Vx) variance. The inheritance of complex traits.com . A complex trait is any trait that does not show simple Mendelian inheritance. genetische variantie. genetische variantie. Kan uitgedrukt worden in variantiecomponenten. Het opsplitsten in genetische en milieuvariantie veronderstelt dat het genotype en milieu ongecorreleerd zijn.o. dan kun je kwantitatieve eigenschappen makkelijker bestuderen. Diabetes is an example of an categorical trait.

Bij een dominant effect varieert de fenotypische waarde door afwijking van de midparent value. Narrow-sense heritability provides a measure of the degree to which the genetic constitution of individuals determines the phenotypes of their offspring.De Marktplaats voor het Kopen en Verkopen van je Studiemateriaal Additive variation is predictably transmitted from parent to offspring. Any deviation of the heterozygote from the midpoint between the two homozygous clasees indicates a degree of dominance of one allele. The additive deviation is know as the breeding value. The genes hat control variation in quantitative (or complex traits) are known as quantitative trait loci (QLTs). de midparent value. . van de geselecteerde ouders. H2 = Va / Vx (Va + Vd + Ve) The genetic deviation (g) of an individual form the population mean is omposed of two parts. The genetic variation for a trait in a population (Vg) can be decomposed into the additive (Va) and the dominance (Vd) variances. When the trait value for the heterozygous class is midway between the two homozygous classes. D>1 overdominantie. dominance variation is not. D is de afwijking van de heterozygoot t. gene action is called additive. het gemiddelde voor die populatie S = Selection differential. D=0 additieve eigenschap.v.o. Zie pagina 684.v.o. Midparent value = het gemiddelde en dat doe je door de twee homozygote genotypen bij elkaar op te tellen en te delen door twee. and it represents the component of an individual’s phenotype that is transmitted to its offspring.Stuvia. See picture pag 671 and pp. its additive deviation (a) and its dominance deviation (d). 681. dus op het gemiddelde. het gemiddelde voor die populatie. H3 kan dus door twee dingen bepaald worden. A = Het verschil tussen de twee homozygote genotype en dat gedeeld door 2.v. H2 = R / S R = Selection response. The additive and dominance effects and their ratio (D/A) provide metrics for quantifying the mode of gene action. The additive variance is the fraction of the genetic variation that is transmitted from parent to parent. door de correlatie tussen de ouders en het nageslacht te bepalen en de ratio van de selectie response t.com . The narrow-sense heritability is the ratio of the additive variance to the phenotypic variance. het gemiddelde afwijking van het nageslacht t.o.v. D=1. Additief wil zeggen dat de heterozygote waarde tussen de twee homozygote waarden in zin.o. dominante eigenschap. D = Afwijking van de heterozygoot van de midparent value. Pag. de gemiddelde afwijking van de ouders (de geselecteerde) t.

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