Human

Genome

Courtesy: National Human Genome Research Institute - Modified from Figure 1.1 Genomes 3 (© Garland Science 2007)
 Human Genome
 The total length is over 3 billion base
pairs
The genome is organized into 22 paired
chromosomes, plus the X chromosome
(one in males, two in females) and, in
males only, one Y chromosome.

These are all large linear DNA molecules

contained within the cell nucleus.

The genome also includes the

mitochondrial DNA, a comparatively
small circular molecule present in each
mitochondrion.

Basic information about these molecules

and their gene content, based on a
reference genome that does not
represent the sequence of any specific
individual, are provided in the following
table.
Figure 1.8a Genomes 3 (© Garland Science 2007)
Figure 1.5 Genomes 3 (© Garland Science 2007)
Figure 1.21 Genomes 3 (© Garland Science 2007)
Figure 2.3 Genomes 3 (© Garland Science 2007)
Figure 2.4b Genomes 3 (© Garland Science 2007)
 Type of DNA ends generated by restriction enzymes

The arrows indicate the cut sites. Phosphate groups attached to the 5' ends
after restriction digestion are indicated in yellow.
 Traditional Cloning Workflow
Using PCR, restriction sites are added to both ends of a dsDNA, which is then digested by the
corresponding REases.

The cleaved DNA can then be ligated to a plasmid vector cleaved by the same or compatible
REases with T4 DNA ligase.

DNA fragments can also be moved from one vector into another by digesting with REases and
ligating to compatible ends of the target vector.
Plasmids
 Plasmids

 A plasmid is a small DNA molecule within a cell that is

physically separated from a chromosomal DNA and can
replicate independently.

 The term was coined by Lederberg and Hays and shortly

discovered by Tatum.

 They are most commonly found in bacteria as small

circular, double-stranded DNA molecules; however,
plasmids are sometimes present
in archaea and eukaryotic organisms.
 Understanding a Plasmid

 Many bacteria have accessory DNA

molecules in addition to their larger
chromosome.

 These molecules, called plasmids, are

extensively used in genetic engineering.

 In order to be useful in labs, these

plasmids need to have an origin or
replication (ori), which enables them to
replciated within a bacterial cell.

 They also need to have multiple

restriction enzyme sites to enable
cutting and pasting of DNA into a
plasmid.
 Plasmids

 Plasmids carry genes that may benefit the survival of

the organism, for example antibiotic resistance.

 While the chromosomes are big and contain all the

essential genetic information for living under normal
conditions, plasmids usually are very small and contain
only additional genes that may be useful to the
organism under certain situations or particular
conditions.

 Artificial plasmids are widely used

as vectors in molecular cloning, serving to drive the
replication of recombinant DNA sequences within host
organisms.