Powerpoint 2 Chapter 5

Question 1

? cleave polypeptides at specific symmetry sites

Answer 1

restrictions enzymes

Question 2

southern blots identify specific ? sequences

Answer 2

DNA

Question 3

northern blots identify specific ? sequences

Answer 3

RNA

Question 4

chemical synthesis of DNA

Answer 4

a deoxyribonucleoside with DMT and BCE attached couples to OH group of resin nucleotide. Elongation continues by deprotection with dichloroacetic acid

Question 5

Sanger sequencing

Answer 5

primer is atached to DNA template to be sequenced and a DNA polymerase is added along with labelled sNTPs and a dideoxy dNTP.

Question 6

PCR

Answer 6

add excess flanking primers of desired DNA sequence. heat to separate strands. cool to anneal primers. synthesize new DNA. heat to seperate. cool to repeat synthesis.

Question 7

vector

Answer 7

a DNA molecule that can replicate autonomously

Question 8

Plasmids

Answer 8

circles of DNA that act as accessory chromosomes in bacteria, can replicate independently.

Question 9

plasmids cut by ? for recombination

Answer 9

restriction enzymes

Question 10

cut plasmids can be annealed using ?

Answer 10

DNA ligase

Question 11

what if DNA is not naturally flanked by appropriate restriction sites?

Answer 11

DNA linker containing restriction site added to ends of DNA

Question 12

insertional inactivation

Answer 12

inserting DNA within a gene to inactivate

Question 13

polylinker

Answer 13

contains many unique restriction sites

Question 14

benefits of bacterial and yeast artificial chromosomes

Answer 14

allow cloning of larger inserts of DNA

Question 15

requirements to clone small DNA fragment in a large DNA collection (2)

Answer 15

specific oligonucleotide probe for gene of interest and a DNA library that can be screened rapidly

Question 16

how is a specific probe obtained?

Answer 16

by synthesizing a oligonucleotide sequence of codons complementary to part of the amino acid sequence of a protein

Question 17

preferred amino acids of Probes

Answer 17

methionine and tryptophan because they only have one codon

Question 18

lambda phage

Answer 18

a virus that is used as a vector for cloning in E. Coli

Question 19

creation of genomic library

Answer 19

genomic dna is fragmented and joined to lambda phages. lambda phages infect E. Coli and replicated. The phages lyse the E. coli host. Sufficiently large number of virus particles to ensure entire genome is represented.

Question 20

how to screen a genomic library for a specific gene?

Answer 20

plate phages of genomic library on lawn of bacteria. Apply nitrocellulose paper to make copy. Add NaOH and labelled probe. Add x-ray film to identify label.

Question 21

oligonucleotide directed mutagenesis

Answer 21

a primer containing a mismatched nucleotide is used to produce a change in DNA sequence and in turn a single amino acid substitution

Question 22

cassette mutagenesis

Answer 22

deletion of gene in a plasmid by cutting with restriction enzymes, purifying the remaining fragment, and adding a new gene (cassette) through ligation to reform the plasmid

Question 23

recombinant HGH replaced HGH isolated from ?

Answer 23

cadavers

Question 24

transcriptome

Answer 24

the entire set of RNA molecules in a population of cells

Question 25

size of genome does/does not reflect number of genes

Answer 25

does not

Question 26

SINES

Answer 26

repetitive DNA elements

Question 27

number of pairs of autosomes in human genome

Answer 27

22

Question 28

changes in gene expression in response to stimuli can be monitored on ?

Answer 28

microarrays/ gene chips

Question 29

microarray

Answer 29

oligonucleotides affixed to suport, labelled cDNA allowed to hybridize to reaveal expression level for each gene, identifiable by known position on microarray

Question 30

episomal

Answer 30

not integrated into bacterial genome

Question 31

rat growth hormone containing DNA can be injected into fertilized mouse egg with ? producing transgenic mouse that produces 500 times mores somatotropin

Answer 31

micropipette

Question 32

Ti plasmids

Answer 32

used to introduce foreign genes into plants

Question 33

types of site-specific double strand breaks

Answer 33

non-homologous end-joining repair, homologous recombination repair (better)

Question 34

non-homologous end-joining repair

Answer 34

error-prone. used for gene disruption or knockout

Question 35

homologous recombination repair

Answer 35

error-free. used for gene replacement (or knockout)

Question 36

RNA intererence

Answer 36

introduce double stranded RNA. cleaved by DICER into 21bp siRNA. siRNA loaded into RISC protein. one strand cleaved and removed. remaining strand recognizes and binds mRNA which it in turn cleaves and releases.

Question 37

(gain/loss) of myostatin leads to bulked up mice

Answer 37

loss

Question 38

? factors control gene expression

Answer 38

transcription

Question 39

engineered transcription factors implementing domains such as ? or ? can be used to activate or repress gene expression

Answer 39

zinc finger or tal effectors

Question 40

Zinc finger nuclease

Answer 40

generated by fusing a zinc finger DNA-binding domain to a DNA-cleavage domain

Question 41

CNBr cleavage

Answer 41

carboxyl side of Tryptophan

Question 42

Trypsin cleavage

Answer 42

carboxyl side of lysine and arginine

Question 43

chymotrypsin cleavage

Answer 43

carboxyl side of tyrosine, tryptophan, phenylalanine, leucine, and methionine

Question 44

carboxypeptidase A cleavage

Answer 44

amino side of C-term Amino acid (excluding arg, lys, pro)