molecular bio lab

Question 1

gene

Answer 1

a length of DNA that codes for a biologically useful product

Question 2

plasmids

Answer 2

extra chromosomal genetic elements

• double stranded circular DNA molecules
• replicate independent of chromosomal (allows for amplification)
• carry non-essential genes
• can be natural or engineered, can carry antibiotic resistant genes

Question 3

pGlO

Answer 3

carries three genes and an ori (origin of replication)

Question 4

GFP

Answer 4

green fluorescent protein, the gene we are cloning

Question 5

araC

Answer 5

a regulatory protein, links GFP expression to presence of arabinose

Question 6

bla/ beta-lacatamase

Answer 6

an enzyme that breaks down ampicilin and makes plasmid-bearing cells antibiotic resistance

Question 7

ori

Answer 7

origin of replication
- a region of plasmid DNA where proteins bind to initiate DNA replication (needed to make more copies of the plasmid in cells)

Question 8

PCR steps

Answer 8

1. denature template strands by heating, strand separate (95 degrees)
2. Cool to let primers anneal; primers bind (55 degrees)
3. allow polymerase to extend to primers; nucleotides add to 3’ end of primers

repeat cycle 25-30 times resulting in a billion fold increase in source DNA

Question 9

genetic engineering

Answer 9

• the ability to move and join pieces of DNA
• purposes: study of gene, purification of protein, alteration of plants, animals
• need enzymes to cut and paste DNA, and a carrier (vector) for DNA

Question 10

Restriction Endonucleases

Answer 10

• bacterial proteins which bind DNA at a specific site and cleave a phosphodiester bond
• recognition site: usually 6 bp and palindromic, protein binds a dimer

Question 11

DNA ligase

Answer 11

• enzyme that makes phosphodiester bonds between pieces of DNA
• normally joins okazaki fragments during DNA replication

Question 12

vectors

Answer 12

• a gene by itself will not replicate on its own within a cell
• must be attached to something with replication controls= could be chromosomal ori or disabled virus

Question 13

cloning

Answer 13

• cutting and pasting allows for recombinant DNA

- cloning is making large amounts of identical copies

Question 14

getting DNA into a cell to clone

Answer 14

transformation- uptake of naked DNA
transjection- use of a viral vector
injection- using a fine glass needle
electroporation- electric shock makes pores
projectiles- gold and tungsten beads covered in DNA

Question 15

agarose gel electrophoresis

Answer 15

• DNA is electronegatively charged so it migrates to positive charge
• smaller fragments of the DNA go through faster and farther