Recombinant DNA Technology and Molecular Cloning

Question 1

It is the process of isolating a specific DNA sequence (like a gene) and creating multiple identical copies of it

Answer 1

Gene cloning or Molecular cloning

Question 2

It involves transferring the DNA of interest into a self-replicating genetic element like a plasmid, which can then be propagated in a host cell

Answer 2

Gene cloning or Molecular cloning

Question 3

named for preventing invasion by foreign DNA by cutting it into pieces

Answer 3

Restriction endonucleases

Question 4

enzymes that cut at sites within the foreign DNA instead of chewing from the ends

Answer 4

Restriction endonucleases

Question 5

by cutting at specfic sites they function as finely honed molecular knives

Answer 5

Restriction endonucleases

Question 6

True or False: Restriction endonucleases are named using the first three letters of their name from the Latin name of their source microorganism

Answer 6

True

Question 7

recognize a specific DNA sequence, cutting only at that sequence

Answer 7

Restriction endonucleases

Question 8

True or False: The restriction endonucleases can recognize 4-bp, 6-bp, and 8-bp sequences

Answer 8

True

Question 9

True or False: The frequency of cuts of Restriction endonucleases increases when the recognition sequence is longer

Answer 9

False, the frequency of cuts lessens

Question 10

Many restriction endonucleases make staggered cuts in the 2 DNA strands which leaves single stranded overhangs called _ that can base pair together briefly

Answer 10

sticky ends

Question 11

occur when the recognition sequence usually displays twofold symmetry, palindromes

Answer 11

Staggered cuts

Question 12

What prevents the restriction endonucleases from cutting up the host DNA

Answer 12

They are paired with methylases

Question 13

enzymes that methylate

Answer 13

methylases

Question 14

If restriction endonucleases and methylases are combined, they form

Answer 14

R-M system

Question 15

small circular pieces of DNA independent of the host chromosome

Answer 15

plasmids

Question 16

joins 2 pieces of plasmid with covalent bonds

Answer 16

DNA ligase

Question 17

function as DNA carriers to allow replication of recombinant DNAs

Answer 17

Vectors

Question 18

Typical experiments uses how many vectors plus a piece of foreign DNA?

Answer 18

one

Question 19

True or False: Foreign DNA has no origin of replication

Answer 19

True

Question 20

site where DNA replication begins

Answer 20

origin of replication

Question 21

Plasmid as vectors

Answer 21

Multiple cloning sites (polylinker)
Origin of Replication
Selectable marker

Question 22

They are those that can yield protein products of the cloned genes

Answer 22

Expression vectors

Question 23

bacterial vectors have a strong promoter and a ribosome binding site near _

Answer 23

ATG codon

Question 24

have a strong promoter and a ribosome binding site near ATG codon

Answer 24

bacterial vectors

Question 26

small piece of DNA which is used to reintroduce the foreign gene of interest into the host cell

Answer 26

cloning vector

Question 27

plasmid that not only introduces a gene of interest into host organism but also aids in the analysis of the gene of interest via relevant protein product expression

Answer 27

Expression vector

Question 28

What is the role of cloning vector?

Answer 28

obtain multiple copies of the foreign gene of interest

Question 29

What is the role of expression vector?

Answer 29

obtian or analyze the gene product, which may be RNA or protein, of the inserted gene of interest

Question 30

What are the different types of cloning vector?

Answer 30

plasmids cosmids phages BACs YACs MACs

Question 31

What are the different type of expression vector?

Answer 31

Only plasmids can be an expression vector

Question 32

What are the features of cloning vector

Answer 32

Origin of Replication Unique restriction sites reporter gene antibiotic resistance site

Question 33

What are the features of expression vectors?

Answer 33

regulatory elements like: enhancers promoters termination sequences transcription initiation site translation initiation site features of cloning vector

Question 34

collection of DNA fragments stored and propagated in a population of microbes through molecular cloning

Answer 34

DNA libraries

Question 35

What are the two types of DNA libraries

Answer 35

Genomic libraries cDNA libraries

Question 36

used to study genes, identify specific DNA sequences, and clone genes of interest

Answer 36

DNA libraries

Question 37

contain fragments of DNA representing the entire genome of an organism

Answer 37

Genomic Libraries

Question 38

can be used to study the organization of the genome, identify regulatory sequences, and clone entire genes

Answer 38

DNA libraries

Question 39

contain DNA sequences that have been reversed transcribed from mRNA

Answer 39

cDNA libraries

Question 40

these libraries are useful for studying gene expression, identifying alternatively spliced transcripts, and cloning expressed genes

Answer 40

cDNA libraries

Question 41

this involves using labeled probes (complementary DNA or RNA) to bind to the target sequence within the library

Answer 41

Hybridization-based screening

Question 42

Clones containing the target sequence are identified by detecting the hybridized probe

Answer 42

Hybridization-based screening

Question 43

this method relies on detecting the protein product of the gene of interest

Answer 43

Expression screening

Question 44

Antibodies can be used to bind to the expressed protein, identifying clones containing the gene.

Answer 44

Expression screening

Question 45

It involves the use of primers specific to the target sequences, PCR can be used to amplify and identify clones containing the target DNA

Answer 45

PCR screening

Question 46

a genetic transfer process by which free DNA is incorporated into a recipient cell and brings about genetic change

Answer 46

Transformation

Question 47

What are the requirements of Transformation?

Answer 47

DNA must be present in extracellular environment Recipient bacteria must be in a state of competence Translocated DNA must be stabilized

Question 48

How can we stabilize translocated DNA as a requirement for transformation?

Answer 48

by integration into the recipient genome by recircularisation (in the case of plamid DNA)

Question 49

ability of a bacterium to take up DNA from the medium

Answer 49

competence

Question 50

cell's ability to take up foreign DNA from its environment

Answer 50

competence

Question 51

True or False: competent cells can only be naturally competent, not made artificially competent

Answer 51

False, it can also be made artificially competent through laboratory techniques

Question 52

induction of competency in cells through chemical means

Answer 52

Artificial competency

Question 54

its traditional method involves incubating bacterial cells in concentrated calcium salt solution

Answer 54

chemcial transformation

Question 55

this solution makes the cell membrane leaky, permeable to the plasmid DNA

Answer 55

concentrated calcium salt solution

Question 56

newer method that uses high voltage to drive the DNA into the cells in process called _

Answer 56

electroporation

Question 57

microprojectile or particle bombardment

Answer 57

Biolistic transformation

Question 58

uses DNA coated tungsten projectiles at high velocity to penetrate cells

Answer 58

Biolistic transformation

Question 59

It is commonly used in yeast, algae, and plants molecular cloning

Answer 59

biolistic transformation

Question 60

What are the two types of Animal Transfection?

Answer 60

calcium phosphate liposomes

Question 61

mix cells with DNA in a phosphate buffer then solution of calcium salt is added to form precipitate

Answer 61

calcium phosphate

Question 62

DNA mixed with lipid to form liposomes, small vesicles with some of the DNA inside

Answer 62

liposomes

Question 63

use of a very fine pipette to inject DNA molecules directly into the nucleus of the cells to be transformed

Answer 63

microinjection

Question 64

involves using the bacterium Agrobacterium tumefaciens to introduce foreign gnees (transgenes) into plant cells, ultimately leading to the integration of these genes into the plant's genome

Answer 64

Agrobacterium-mediated transformation

Question 65

What does A. tumefaciens contain?

Answer 65

T-DNA

Question 66

derived from a plasmid known as tumor-inducing (Ti)

Answer 66

T-DNA

Question 67

comes from bacteria that cause plant tumors called crown galls

Answer 67

Ti plasmid

Question 68

plant tumors

Answer 68

crown galls

Question 69

T-DNA genes direct the synthesis of unusual organic acids, _ which can serve as an energy source to the infecting bacteria but are useless to the plant

Answer 69

opines

Question 70

allow scientists to distinguish between transformed and nontransformed products

Answer 70

selectable markers

Question 71

It is important to eliminate nontransformed cells using negative or positive selection techniques

Answer 71

selectable markers

Question 72

It involves marker gene (antibiotic resistant marker)

Answer 72

negative selection

Question 73

only cells that express resistant gene survive

Answer 73

Negative selection

Question 74

What is an example of Negative selection?

Answer 74

hygromycin-resistance gene

Question 75

What is an example of positive selection?

Answer 75

marker gene encoding PMI