Genetic Engineering

Question 1

Genetic Engineering

Answer 1

the direct manipulation of an organism’s genes

Question 2

Polymerase Chain Reaction (PCR)

Answer 2

1. When DNA is heated to 95o the strands separate
2. At 72o or cooler the bases pair up
3. A DNA polymerase enzyme will “fill in” the missing bases

Question 3

PCR Limitations

Answer 3

• Information about the sequence of the target DNA must be known
• Contamination
  o Non-specific DNA strands can be easily amplified
• Controls
  o Negative controls needed (i.e. with no template DNA)
• Primer optimisation
  o Best temperatures for annealing, primer length, concentrations

Question 4

DNA Technology - Vectors

Answer 4

Vectors
DNA carrier molecules that transfer and replicate (clone) inserted DNA fragments

Characteristics of vectors:
1. Able to independently replicate itself and any DNA fragments it carries
2. Contain recognition sequences to allow insertion of fragments
3. Carry a selectable marker to distinguish cells that have adopted vectors
4. Easily recoverable from the host cell

Question 5

Plasmids

Answer 5

A plasmid is a small DNA
molecule within a cell that is
physically separated from
chromosomal DNA and can
replicate independently

Question 6

rDNA Technology

Answer 6

1. DNA to be cloned is purified
   from cells or tissues
2. Gene isolated from DNA template either via PCR (for amplification) or
   Restriction enzymes (to specifically cut out gene)
3. The fragments produced are joined to other DNA molecules called vectors (carrier molecules). A vector joined to a DNA fragment is a recombinant DNA (rDNA) molecule
4. The recombinant DNA molecule is transferred to a host cell
5. As host cells replicate, the recombinant DNA molecules within them are passed on to
   all their progeny, creating a transgenic population
6. The cloned DNA can then be
   recovered from the host cells,
   purified and analysed
7. The cloned DNA is then transcribed and translated, and the encoded gene product isolated and used (research or commercial)