Week 7 Bacterial Isolation

Please answer the following as brief as possible and in your OWN words.

1. Explain the term selective and non selective media with proper examples (5 points).

 A selective media is one that specifically promotes the proliferation of one or more specific
microorganisms while suppressing the growth of other microbes. MacConkey agar with the
existence of bile salt is one example; it is a medium that inhibits the development of Gram-
positive bacteria while permitting the growth of Gram-negative bacteria. Nonselective media
doesn’t have a selection agent which means it permits different microorganisms to grow, it is
also a simple media created to grow bacteria. An example of nonselective media is blood agar.

2. In short paragraph, tell us the story of Typhoid Mary? (5 points).

 Cooking for a family in New York was Mary Mallon's job. Despite showing no symptoms, Mary
eventually had typhoid. Between 1900 and 1907, Mary employed at least eight homes, all of
which had suffered serious typhoid damage. Mary, an Irish woman with a large build, was the
most frequently affected family member, according to a doctor's observation in 1907. After
being imprisoned in her home for three years, she was finally given permission to depart, but
only if she vowed never to again prepare meals. Immediately afterward, she went back to her
profession as a cook and spread the disease for a further five years. She lived her entire life in
isolation on an island after causing a major pandemic in 1915.

3. What is the significance of RNA extraction? (10 points).

 DNA in a cell includes genetic code that is transcribed into the proteins that determine the
structure and function of the cell. This technique is mostly due to the importance of RNA
extraction.

Week 8 Nuclei Acid Isolation

Please enumerate and explain the different steps in the nucleic acid isolation as brief as possible and in
your OWN words (20 points).

 Step 1: Cell Lysis - The first procedure in isolating nucleic acids is cell lysis, or rupturing the cell
wall and/or membrane to release the genetic material. By using a lysis buffer, rotor
homogenizer, bead mill, freeze-thaw cycles, or sonication, this is accomplished.
 Step 2: Precipitation – The DNA is already outside the cell after lysis, but it is still diversified with
the other elements. The following step will use sodium ions to balance the DNA molecules'
negative charges before adding alcohol to precipitate the DNA out of the aqueous solution and
separate it from other debris. The next step is purification because additional cellular debris
such as protein and lipids that may obstruct analysis are still present after these.
 Step 3: Purification – To get rid of any additional or lingering unwanted particles, alcohol is
applied as a rinse. The DNA is then frequently redissolved in water or an eluting solution for
easier handling and storage.
Week 9 RNA Extraction

Please answer the following as brief as possible and in your OWN words.

1. What is the main purpose of nucleic acid isolation? (5 points).

 By lysing or disassembling the cell wall and removing as many lipids and proteins as possible, the
main goal of nucleic acid isolation procedures is to produce pure and extremely productive DNA
and RNA.

2. How do we make sure that we get a high purity of nucleic acid during nucleic acid isolation? (5 points).

 The lysis, precipitation, and purification processes—which are the three fundamental phases in
isolation—must be carried out if we want to ensure that the nucleic acid we obtain during
nucleic acid isolation is of a high purity. The purification process is the most crucial of the three
processes. If high yield and purity of DNA are needed for the separated DNA's subsequent use, it
is carried out.

3. Why do we need to isolate RNA? (5 points).

 Using the genetic codons found on messenger RNA, proteins are synthesised. Which genes are
expressed depends on the types and quantities of RNA that are present in a cell. Isolated RNA
performs well in a number of downstream applications, including primer extension, reverse
transcription, RNA protection tests, next-generation sequencing, and qPCR microarrays. RNA
extraction aims to create high-quality, extremely pure RNA for usage from biological sources.

4. Enumerate and explain briefly the experiments that nucleic acid can be used after isolation (5 points).

 In the lab, a process known as polymerase chain reaction (PCR) is used to produce several copies
of a certain DNA fragment from a substance that only contains extremely small amounts of that
DNA. These DNA fragments can be amplified and identified using the polymerase chain reaction.
 DNA and RNA fragments are separated, recognized, and purified using the molecular biology
technique known as electrophoresis based on size and charge.

Week 10 qPCR (Gene Expression Unit)

Please answer the following as brief as possible and in your own words.

1. Give at least 5 applications of qPCR (10 points).

a. Genotyping. When employing molecular probes for genotyping, real-time PCR/ qPCR provides a high
throughput option for quick and precise results.
b. Food Safety. QPCR, which assists in discovering both incidental and intentional adulterations of food
by biological pollutants, can find targeted DNA segments.
c. Gene Expression Analysis. For measuring gene expression, molecular biologists typically employ the
quick, easy, and inexpensive qPCR method. Using endogenous controls such as reference genes, qPCR is
used to assess the relative expression of a target gene.
d. Cancer Phenotyping. The identification and quantification of genes and gene expression in a variety
of sample sources is made possible by qPCR molecular methods. These techniques make it possible to
identify potential cancer biomarkers with speed and accuracy.
e. Detection of GMOs in food. A qPCR-based approach is the most reliable choice for GMO identification
because to its great sensitivity and specificity.

2. Explain the benefits of using SYBR green to quantify DNA in qPCR and gel electrophoresis as compared
to the use of ethidium bromide (10 points).

 With SYBR Green, qPCR primer designs do not need to incorporate fluorescent reporter systems,
fluorescently-tagged probes, or beacons that are particular to a target sequence. SYBR Green
qPCR formulations are suitable for all real-time PCR methods, including fast PCR. SYBR Safe DNA
Gel Stain was developed as an ethidium bromide alternative that is safer. SYBR Safe DNA Gel
Stain was the subject of numerous tests carried out by independent, authorized testing
agencies, which found minimal to no genotoxicity and no acute toxicity.

Week 11 Stem Cell Technology (Gene Regulation & NGS)

Please answer the following as brief as possible and in your own words.

1. Give at least 5 applications of stem cell technology. (10 points)

a. Therapy for Blood Disorders. Nowadays, leukemia, sickle cell disease, and other immunodeficiency
problems are often treated with adult hematopoietic stem cells.
b. Treatment for Brain Disorders. When brain cells are destroyed, muscles cannot be controlled.
Scientists may be able to restore damaged brain tissue using stem cells. As a result, the specialized brain
cells that prevent uncontrollable muscle movements can reappear.
c. Regeneration of Tissues. In the absence of organ donors, stem cells may be used by researchers to
create a certain tissue type or organ by being instructed to develop in a certain manner.
d. Treatment for Cell Deficiency. These new cells may be able to repair cardiac injury by reintroducing
healthy tissue into the heart. Patients with type I diabetes may receive pancreatic cells from their own
immune systems to replace any lost or destroyed insulin-producing cells.
e. Treatment for Heart Disease. Only two weeks after the stem cells were inserted, networks of blood-
perfused arteries emerged. These artificial blood vessels were of the same caliber as the native ones in
the area.

2. If you were to use stem cell technology to cure only ONE disease, what is this and WHY? (10 points).

 Since coronary heart disease continues to be the biggest cause of mortality in our country, if I
could only use stem cell technology to cure one illness, I would choose that one. Heart disease is
the most common illness in the entire world. This condition may lead to the patient's death.
Patients between the ages of 20 and 30 are experiencing an increase in heart attacks. Younger
people are just as susceptible to heart attacks as older adults are. Age is not a factor.