recombinant DNA techh

Question 1

steps of setting up PCR (basic)

Answer 1

rna purification
then reverse trancription to get it to DNA
then PCR

> denature, anneal, DNA synthesis

Question 2

which procedure needs a primer?
A cDNA sythesis
B genration of transgenic plants (Ti plasmid)
C ligation of DNA fragment
D restriction enzyme digestion
E transcription in vitro

Answer 2

cDNA synthesis
> reverse transcriptase uses oligodT primer
> pairs with polyAAA tail and reverse transcriptase = hybrid

Question 3

which enzyme is used to join 2 dna fragments with compatible sticky ends
A rna polymerase
B dna helicase
C dna gyrase
D Ddel
E dna ligase

Answer 3

dna ligase!!

it ligates = joining by covalent bonds and can sythesise new phosphodiester bonds with ATP.

Question 4

AquAdvantage Salmon are a transgenic animal that were made to produce more growth hormone. This was done by…
A Increasing their production of growth hormone stimulating hormone
B Adding the promoter for an antifreeze gene of another species
C Increasing the amount of somatotrophs present in the salmon’s anterior pituitary
D Adding a gene that prevents the decreases somatostatin release

Answer 4

OPTION B ..
AquAdvantage Salmon were made by by fusing the promoter for an antifreeze gene from an Ocean Pout, to the growth hormone cDNA from a salmoN
> ANTIFREEZE gene is present in all tissues so now growth hormone is present in all tissues instead of just pituitary gland

Question 5

PCR is…
A The targeted amplification of a specific DNA sequence
B The amplification of a random DNA sequence
C The targeted amplification of a specific RNA sequence
D The amplification of a random RNA sequence

Answer 5

OPTION A
PCR is the targeted amplification of a specific DNA sequence by a billion fold. It is made specific by the synthesis of oligonucleotide single stranded DNA primers.
> does not work on RNA as taq polymerase doesnt recognise uracil

Question 6

A transgenic animal is one that…
Has been cloned

Has exogenous DNA inserted into its genome

Has had a sequence of DNA removed from its genome

Has had its DNA undergo PCR

Answer 6

OPTION B

Question 7

what is an advantage of using recombinant DNA tech in medicine - pregancy?

Answer 7

you can non inasively analyse foetal DNA

> also applications in foresnics using only a very small amount of blood

Question 8

One stranded of a molecule of double stranded DNA has the sequence: 5ʹ-CGAA-3ʹ.  What is the sequence of the complementary strand? 
5ʹ-AAGC-3
5ʹ-ATCC-3ʹ
5ʹ-CCTA-3
5ʹ-GCTT-3
5ʹ-TTCG-3

Answer 8

OPTION E
as we go in the 5-3 direction
> dna is anti parallel

Question 9

name some requirements for plasmids

Answer 9

UNIQUE single site for restriction enzymes

selectable marker that allows cells with plasmid to be distinguished from cells wthout it

Question 10

how can we distinguish between recombinant plasmids and reformed plasmid (without the dna gene

Answer 10

the restriction enzyme site is in LacZ gene so recombinsnt plasmid cant make b-galactosidase so will not give a coloured dye.

Question 11

how do the shine dalgamo and kozac sequences differ?

Answer 11

dalgamo pairs up with ribosome at binding site

kozac overlaps with initiation codon and needed for efficient translation

Question 12

how can we overcome issues with expressing eukaryotic genes in E.coli?

Answer 12

use cDNA library/ mrna transcript which lack introns

use expression vectors that contain prokary promotoer, terminators and ribosomal binding site!

Question 13

which enzyme is involved in PCR?
A oligio-dT primer
B dna ligae
C rna polymerase
D Taq DNA polymerase
E Taq RNA polymerase

Answer 13

OPTION D

extracted from heat resistant bacteria so heat stable!

Question 14

how does DNA profiling work?

Answer 14

uses short tandem repeat (2-6bp, 1-50 repeat)

> number of repeats vary in indiviuals so analyse lots of these STR

Question 15

how is real-time PCR different to PCR

Answer 15

it measures the accumulation of amplified DNA after each cycle using a fluorescent probe

Question 16

transgenic goat facts

Answer 16

we can get antithrombin (prevent blood clotting) from goats milk
> fuse the anththrombin gene with the milk protein gene

Question 17

what ways can dna be damaged?

Answer 17

uv , gamma or x-rays

Question 18

how can dna be repaired

Answer 18

homology directed repair - only in Sphase using sister chromatid as template
non-homologous end joining - any time but error prone as a INDEL is always added

Question 19

which of the following is true of PCR?
A denaturation at 72°C
B primers 5-10bp in length
C uses DNA polymerase from mouse cells 
D after 20 cycles, DNA amplified by 2^20
E synthesis at 55°C

Answer 19

OPTION D IS TRUE

denatuation 95, anneal 45, synthesis 72
dnapolymerase from thermophiles
primers are 20bp length!

Question 20

Which of the following is NOT true of the polymerase chain reaction (PCR)?
A DNA denaturation 95 degrees
B DNA synthesis 72 degrees
C After 20 cycles of PCR the amount of the DNA
amplified increased 20x
D needs dATP dCTP dGTP dTTP
E needs primers complementary to ends of DNA

Answer 20

OPTION C

dna increase by 2 to power of 20!!

Question 21

which of the following NOT needed for Sanger sequencing
A DNA ligase
B Dideoxynucleoside triphosphate
C Deoxynucleoside triphosphate
D A primer
E DNA polymerase

Answer 21

OPTION A
Dideoxy is chain-terminating as it lacks a hydroxyl
> for a single gene/DNA fragment

Question 22

which best describes reverse transcription
A mRNA as template synthesises complementry strand of DNA
B mRNA as template synthesises complementry strand of RNA
C protein as template synthesises mRNA encoding the protein
D mRNA as template synthesises mRNA encoding the protein

Answer 22

OPTION A

Using an mRNA molecule as a template, it synthesises a complementary strand of DNA

Question 23

restrction enzymes can be obtained from
A bacteria
B fungi
C algae
D virus
E plants

Answer 23

BACTERIA! restrction endonucleaese found in their immune system

Question 24

how can we overcome expressing eukaryotic genes in bacterial cells?

Answer 24

use EXPRESSION VECTORS have
> prokary promotion and termination sequence
> ribosomal binding site

Question 25

which blotting uses a labelled DNA/RNA probe but detects RNA?
A northern blot
B southern blot
C eastern blot
D western blot

Answer 25

OPTION A

noRthen blot

Question 26

why do we use Ti (tumor inducing) plasmid to make transgenic plants?

Answer 26

during an infection it becomes intergrated into a plants genome
> aslo need a VIR REGION to transfer the T-region of DNA into chromosome

Question 27

What is the name of the process that uses a fluorescent oligonucleotide probe?
DNA profiling
Polymerase chain reaction
Fluorescence profiling
Real time polymerase chain reaction
Transgenic recombination

Answer 27

OPTION D

Real time PCR (qPCR) uses a fluorescent probe so that accumulation of product can be measured during the reaction.

Question 28

Eco RI is an example of a restriction enzyme that recognises 6bp target sites. After cleavage, what kinds of ends does it leave?
A Blunt ends
B Overt ends
C Sticky ends
D Static ends
E Bowed ends

Answer 28

OPTION C
Eco RI recognises 6bp target sites and cleaves to leave sticky ends.
Hae II recognises 4bp target sites and cleaves to leave blunt ends.