Question

1. A researcher wants to clone genes using recombinant gene technology as part of a genome sequencing project. There are eight steps below. First, cross out the two steps that are not part of this process. Then, put the remaining six steps in the correct order.

A Insert recombinant plasmid into bacterium.

B Cut the genome into fragments using restriction enzymes.

C Determine activity using laser.

D Use DNA Ligase.

E Make cDNA using reverse transcriptase.

F Grow bacteria and add clone to genomic library.

G Cut bacterial plasmids with restriction enzymes.

H Combine DNA fragments and cut plasmid DNA.

Answer 1

The following forms the reactions

1. Cut the bacterial Plasmid with a restriction enzyme for further reaction or cloning purpose

2. Cut the genome into fragments using restriction enzyme

3. Use DNA ligase

4.Combine DNA fragment and Cut plasmid DNA

5.Insert the recombinant plasmid into the bacterium

6. Grow bacteria and add clones into genome library

Following depicts the Plasmid Gene Technology

Bacterial cell lacz gene (lactose breakdown) Human cell Isolate plasmid DNA and human DNA. Restriction site ampf gene Bacterial (ampicillin plasmid resistance) Cut both DNA samples with the same restriction enzyme. C Gene of interest Sticky ends Human DNA fragments OC Mix the DNAs; they join by base pairing. The products are recombinant plasmids and many nonrecombinant plasmids. OO Recombinant DNA plasmids Introduce the DNA into bacterial cells that have a mutation in their own lacz gene. Recombinant bacteria Plate the bacteria on agar containing ampicillin and X-gal. Incubate until colonies grow. Colony carrying non- recombinant plasmid with intact lacZ gene Colony carrying recombinant plasmid with disrupted lacZ gene Bacterial clone