Foreign

Gene of interest

This is NOT BREAD!

What? Who am I? I dont know anymore.

6 Questions (30 seconds each) 1 Short Essay (7 minutes) 1) Which of the following exists as DNA surrounded by a protein coat? a) Retrovirus b) Virus c) Eukaryotic cell d) Prokaryotic cell e) Ampicillin

2) Which enzyme is used to make multiple copies of genes in the polymerase chain reaction (PCR)? A. restriction enzyme B. DNA ligase C. reverse transcriptase D. RNA polymerase E. DNA polymerase

3) __________ separates molecules by movement due to size and electrical charge. A. restriction enzymes B. gene cloning C. DNA ligase D. gel electrophoresis E. reverse transcriptase

4) In a nucleosome, the DNA is wrapped around

A) polymerase molecules. B) ribosomes. C) histones. D) the nucleolus.

5) In genetic engineering, DNA ligase is used for which of the following purposes? a. To act as a probe for locating clone genes b. To create breaks in DNA in order to allow foreign DNA fragments to be inserted c. To seal up nicks created in newly created recombinant DNA d. To ensure that sticky ends of like DNA fragments do not re-anneal

6) Restriction enzymes are generally used in the laboratory for which of the following reasons? a. Restricting the replication of DNA b. Restricting the transcription of DNA c. Restricting the translation of mRNA d. Cutting DNA molecules at specific locations e. Cutting DNA into manageable sizes for manipulation

Short essay: Describe PCR and explain the contribution of this procedure. (4 points)

THIS IS AN AGAR PLATE CONTAINING AMPICILLIN

(RIGHT) The E. coli strain DH5a containing the ampicillin resistant pAMP plasmid is able to grow on an agar plate containing ampicillin. (LEFT) The same strain without the pAMP plasmid cannot grow on the ampicillin plate.

MIX MIX MIX! MIX MIX MIX! Lalalallalaa.

Ligated cloning vectors! Woot!

So bacteria may take up plasmid with or without the insert, or may not take up any plasmids at all

How do we pick the colony that has the foreign DNA? Bacteria are grown on media containing ampicillin and X-gal, a chemical that is metabolized by LacZ gene.

Ampicillin kills bacteria without plasmid. (because it does not have the ampR gene Plasmids lacking the foreign insert have an intact lacZ gene and are able to metabolize X-gal which then releases an indicator dye- turning the colony blue Plasmids with an insert with a disrupted lacZ gene will have the foreign DNA in place of it, thus cannot metabolize X-galthus, does not produce blue.. And are WHITE! Ta-da. Done. Finito.

Here is the slide again. Absorb. Absorb. Copy. Label. Color!

Is a genetic variation that can be detected by enzymatic disgestion: CLICK

CLICK