Biotechnology (BIT4001) Plant Biotechnology Tutorial

Tevoney Dawkins

1701707

1. Draw a flow diagram to highlight the general steps involved creating a transgenic plant.
Enzyme restriction of gene of interest ( restriction enzymes BamHI and EcoR1)

Restriction digest of the chosen plasmid vector (typical same as used for digest of gene)

Ligation of DNA products and plasmid vector (done using DNA ligase)

Transformation (electroporation and gene gun can be used)

Growth on agar plates ( callus formation, synthetic seed formation)

2. For each step of the flow diagram above, describe two techniques that can be used to
accomplish it.
3. Briefly describe the strategies used to produce
a. FLAVR SAVR tomato
This is done using gene silencing technology where the PG gene produces an enzyme that is responsible
for the softening of tomato, the sense mRNA which is typically expressed by the plant. The technique
uses an antisense gene which is introduced into transgenic crops. the transgenic crops produce both the
sense and antisense mRNA which are complementary to each other and bind thus reducing the
expression of the PG gene leading to decreased production of the enzyme

b. Virus Resistant Crops

This can be done by using a pathogen derived method where a part of the virus affects the plant. The
coat protein of the virus is transcribed to DNA and then cloned into a plasmid. The gene is then
sequenced and a chimeric gene is produced that is transformed into the plant. The chimeric gene will
have a silencing effect on the regular coat protein of the virus thus negating the expression of the virus.
c. Golden Rice
d. Bt corn
Made using the Bt toxin from bacillus thuringiensis. The gene for the toxin is linked to a promoter to
produce the toxin in all plant cells then is transformed into plants using agrobacterium tumaficiens.

4. Define the following terms

a. Gene silencing
The process of inhibiting the expression of a gene, reducing or preventing its protein production. This
process occurs naturally within cells and can also be induced experimentally. There are several methods
of gene silencing, but two primary mechanisms are RNA interference (RNAi) and DNA methylation.
b. Gene stacking

Gene stacking refers to the process of incorporating multiple genes of interest into an organism's
genome. This technique is often used in genetic engineering or biotechnology to introduce several
desirable traits or characteristics into a single organism.

c. Antisense RNA

Antisense RNA refers to a type of RNA molecule that is complementary to a specific messenger RNA
(mRNA) sequence. This complementary RNA sequence can bind to the target mRNA, forming a double-
stranded RNA structure. This binding can inhibit the expression of the gene encoded by the target mRNA
in a process called gene silencing.

5. Outline three negative impacts of genetically modified crops and give a specific example of each
impact.
6. Outline three positive impacts of genetically modified crops and give a specific example of each
impact.
1. Longer shelf life (Flavr Savr tomato)
2. More nutritious variants ( golden rice)
3. insect resistance ( Bt corn)