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MOLBIO OFFSEM MIDTERM > molecular cloning > Flashcards

molecular cloning Flashcards

1
Q

group of identical cells or organisms which is a product of any cloning experiment

A

clone

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2
Q

some plants can be cloned simple by taking ________ (greek: _______, meaning twig)

A

cutting ; klon

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3
Q

others can be clone by growing whole plants from single cells collected from ___________

A

one plant

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4
Q

To clone a gene one must insert it into a _______ that can carry the gene into a host cell and ensure that it will replicate there.

A

vector

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5
Q

________ - produced clones of identical frogs by transplanting nuclei from a single frog embryo
to many enucleate eggs.

A

John Gurdon

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6
Q

__________ - was cloned in scotland in 1997 using an enucleate egg and nucleus from an adult sheep mammary gland.

A

Dolly the sheep

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7
Q

__________, ________, and _________ - performed first cloning experiment in 1973

A

Stanley Cohen, Herbert Boyer, and Colleagues

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8
Q

(ROLE OF RESTRICTION ENDONUCLEASES)

_________ - depended on invaluable enzymes
called restriction endonucleases.

A

Cohen and Boyer

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9
Q

___________– discovered restriction endonucleases in E. coli in the late 1960’s.

A

Stewart Linn and Werner Arber

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10
Q

____________ - discovered an enzyme from haemophilus influenzae strain Rd, showing specifity in cutting DNA.

A

Hamilton Smith

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11
Q

___________ - usually carried out by cutting the vector and the DNA to be inserted with the same restriction endonucleases to endow them with the same “STICKY ENDS”

A

INSERTION

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12
Q

5 TYPES OF VECTORS:

A

Plasmid vs Vectors

Phages vs Vectors

Cosmids

Phagesmides

Eukaryotic vector and very high pecific vector

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13
Q

Vectors for cloning in bacteria come in two major
types: ________ and _________

A

PLASMID AND PHAGES.

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14
Q

________ and the ________ - among the
plasmid cloning vectors.

A

pBR322 and the pUC plasmids

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15
Q

__________ - is convenient with the pUC plasmids
and pBS phagemids. these vectors have an
ampicilin resistance gene and a multiple cloning
site that interrups a partial b-galactosidase gene
whose product is easily detected with a color test.

A

Screening

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16
Q

________ - are ampicilin- resistant and do
not make active b-galactoside.

A

Desired clones

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17
Q

Two kinds of phage vectors-especially popular as cloning vectors:

A
  1. Lambda
  2. M13 Phages
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18
Q

Two kinds of phage vectors-especially popular as cloning vectors:

  1. __________ - has certain nonessential genes removed to make room for insects. in some of these engineered phages inserts up to 20 kb in length can be accommodated.
A

(lambda)

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19
Q

Two kinds of phage vectors-especially popular as cloning vectors:

  1. ________ - have the convenience of a
    multiple cloning region and producing single
    stranded recombinant DNA which can be used
    for DNA sequencing and for site directed
    mutagens.
A

M13 phages

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20
Q

(IDENTIFYING A SPECIFIC CLONE WITH SPECIFIC PROBE)

2 DIFFERENT KINDS WIDELY USED:

A
  1. Polynucleotides (oligonucleotides) and antibodies
  2. Polynucleotides Probes
21
Q

Specific clones can be identified using _____________ that bind to the gene itself.

knowing the amino acid sequence of a gene product, one can design a set of __________ that encode part of this amino acid sequence.

this can be one of the quickest and most accurate means of identifying a particular _______.

A

polynucleotide probes ; oligonucleotides ; clone

22
Q

_______ - short for complementary DNA or copy DNA, is a
DNA copy of an RNA, usually an mRNA.

A

Cdna

23
Q

________ - a set of clones representing as many as
possible of the mRNA’s in a given cell type at a given time.

A

cDNA library

24
Q

___________ - a clone containing a DNA copy of just one mRNA.

A

Particular cDNA

25
Q

___________ - is like any other DNA synthesizing
enzyme that cannot initiate DNA synthesis without a
primer

A

Reverse transcriptase

26
Q

_____________ - a simultaneous removal of DNA ahead
nick (a single stranded DNA break) and synthesis of DNA behind the nick

A

Nick translation

27
Q

_____________ - usually used for nick translation.

A

E. coli DNA polymerase I

28
Q

(RAPID AMPLIFICATION OF CNDA ENDS)

______ - a procedure used to fill in the missing
pieces of a DNA.

A

RACE

29
Q

________ - illustrates the technique for
filling in the 5-end of a cDNA the usual
problem

A

5- RACE

30
Q

______ - an analogous technique used
to fill in a missing 3- end of a cDNA.

A

3- RACE

31
Q

(POLYMERASE CHAIN REACTION (PCR))

a newer technique that also yield a DNA
fragment for cloning and is especially useful for
cloning cDNA’s

A

Standard PCR

32
Q

Standard PCR amplifies a region of DNA between two
_______________.

A

predetermined sites

33
Q

(POLYMERASE CHAIN REACTION (PCR))

each cycle of PCR double the number of copies
of the ___________ until a large quantity has
been made.

A

amplified DNA

34
Q

_________ and _______ invented PCR in 1980’s

A

Kary Mullis and his colleagues

35
Q
  1. Using RT-PCR in cDNA cloning

__________ a cDNA from just one mRNA whose
sequence is known.

A

RT-PCR cloning

36
Q

DIFFERENCE BETWEEN PCR AND RT-PCR

A

RT-PCR starts with an mRNA instead of a double stranded DNA.

37
Q

_______ can be used to generate a cDNA from a single type of mRNA, but the sequence of the mRNA must be known is the
primers for the PCR step can be designed. ________ sequences can be placed on the PCR primers, so these sites appear at the ends of the cDNA. this makes it easy to cleave
them and then to ligate the _____ into a vector.

A

RT-PCR ; restriction site ; cDNA

38
Q

a way of quantifying the amplification of a DNA as it occurs in real time

A

REAL TIME PCR

39
Q

____________ keeps track of the progress of PCR by monitoring the degradation of a reporter probe hybridized to the strand
complementary to the forward primer. as this probe is degraded, a ______ increase, and this increase can be measured in real time in a __________.

A

Real-time PCR ; fluorescence ; fluorimeter

40
Q

designed to yield the protein product of a cloned gene, usually in the greatest amount possible.

A

Expression vector

41
Q

Expression vector

  • to optimize expression, bacterial expression vectors provide strong _________ and __________ binding sites that would be missing from cloned eukaryotic genes.
A

bacterial promoters ; bacterial ribosomes

42
Q

most cloning vector are inducible to avoid premature overproduction of a foreign product that could poison the bacterial host cells.

A

Inducible expression vector

43
Q

frequently produce fusion proteins, which can often be isolated quickly and
easily.

A

Expression vector that produces fusion protein

44
Q

have the advantages that the protein products are usually soluble, and these
products are modified in a eukaryotic manner.

A

Eukaryotic expression system

45
Q

Other eukaryotic vectors:

A
  1. yeast artificial chromosomes (YACs) bacterial artificial chromosomes (BACs)
    and P1 phage artificial chromosomes (PACs)
  2. Ti plasmid
46
Q

capable of accepting huge
chunks of foreign DNA and are use in large sequencing programs such as the
human genome project.

A

yeast artificial chromosomes (YACs) bacterial artificial chromosomes (BACs)
and P1 phage artificial chromosomes (PACs)

47
Q

another important eukaryotic vector which can transport foreign genes into plant cells and ensures their replication.

A

Ti plasmid

48
Q

cloned genes can also be transferred to plants,
using plant vector such as the Ti plasmid. this
procedure can alter the plants characteristics.

A

Using Ti plasmid to transfer genes to plants

MOLBIO OFFSEM MIDTERM (9 decks)

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