BTEC 1322 LECTURE 11 Recombinant DNA + CLONING

Where are restriction enzymes primarily located?
A) Plants and animals
B) Bacteria and archaea
C) Viruses and fungi
D) Human cell

B) Bacteria and archaea

What precise DNA sequences do restriction enzymes target?

A) Promoter regions

B) Introns

C) Exons

D) Recognition sites or restriction sites

D) Recognition sites or restriction sites

What defines a palindromic sequence in DNA?

A) Sequences with no symmetry

B) Sequences that read the same forward and backward

C) Sequences with overhanging ends

D) Sequences with blunt ends

B) Sequences that read the same forward and backward

What is the term for DNA ends characterized by overhanging, single-stranded regions?

A) Blunt ends

B) Sticky ends

C) Linear ends

D) Palindromic ends

B) Sticky ends

What is the outcome when sticky ends from different DNA fragments, produced by a restriction enzyme, interact?
A) Form a palindrome
B) Generate blunt ends
C) Base-pair with each other
D) Become circular DNA

C) Base-pair with each other

What happens when HindIII and KpnI act on a plasmid containing their recognition sites?

A) They create circular DNA fragments.

B) They generate linearized DNA fragments with blunt ends.

C) They cleave the plasmid DNA, producing linearized DNA fragments with sticky ends

D) They have no effect on the plasmid

C) They cleave the plasmid DNA, producing linearized DNA fragments with sticky ends

What is the purpose of DNA ligase in molecular biology?

A) To amplify DNA segments

B) To denature DNA

C) To cleave DNA at specific sites

D) To join DNA fragments with compatible sticky ends

D) To join DNA fragments with compatible sticky ends

What type of bond does DNA ligase catalyze the formation of?

A) Hydrogen bonds
B) Peptide bonds
C) Phosphodiester bonds
D) Glycosidic bonds

C) Phosphodiester bonds

What is the result of the annealing of complementary sticky ends of the plasmid and insert?

A) Denaturation of DNA
B) Formation of linear DNA fragments
C) Creation of a circular recombinant plasmid
D) Generation of blunt ends

C) Creation of a circular recombinant plasmid

How is the recombinant plasmid introduced into bacterial cells?

A) Through a process called transcription

B) By using a centrifuge

C) Via a procedure called transformation

D) Through PCR amplification

C) Via a procedure called transformation

In the process of preparing recombinant DNA, what crucial function do bacterial cells serve?

A) Breaking down the DNA segments
B) Denaturing the DNA
C) Replicating the recombinant plasmid
D) Inducing mutations in the DNA

C) Replicating the recombinant plasmid

How do plasmids replicate in relation to the bacterial chromosome?

A) They replicate in conjunction with the bacterial chromosome.

B) They rely on the bacterial chromosome for replication.

C) They replicate independently of the bacterial chromosome.

D) They do not undergo replication

C) They replicate independently of the bacterial chromosome.

What is one of the primary uses of PBR322 or pUC19 in molecular biology research?

a) Protein synthesis
b) RNA sequencing
c) DNA cloning and gene expression studies
d) Antibiotic production

c) DNA cloning and gene expression studies

What is the size of PBR322 in base pairs (bp)?

a) Approximately 10,000 bp

b) Approximately 1,000 bp

c) Approximately 20,000 bp

d) Approximately 4,361 bp

d) Approximately 4,361 bp

What are the antibiotic resistance MARKERS present in PBR322?
a) Kanamycin resistance

b) Ampicillin resistance and tetracycline resistance

c) Chloramphenicol resistance

d) Streptomycin resistance

b) Ampicillin resistance and tetracycline resistance

What is the function of the polylinker in PBR322 orpUC19/19?

a) It encodes a protein for DNA replication.

b) It contains multiple unique restriction enzyme recognition sites.

c) It serves as a selectable marker for plasmid-containing cells.

d) It facilitates transcription of genes

b) It contains multiple unique restriction enzyme recognition sites.

How does the size of pUC19 compare to PBR322?
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a) pUC19 is larger than PBR322.

b) pUC19 is considerably smaller than PBR322.

c) pUC19 is approximately the same size as PBR322.

d) pUC19 and PBR322 are unrelated in size.

b) pUC19 is considerably smaller than PBR322.

What selectable marker is present in pUC19?

a) Kanamycin resistance

b) Ampicillin resistance

c) Tetracycline resistance

d) Chloramphenicol resistance

b) Ampicillin resistance

Why is the presence of the ampicillin resistance gene important in pUC19?

a. It allows bacteria to produce ampicillin.

b. It helps in RNA synthesis.

c. It serves as a selectable marker for plasmid-containing cells.

d. It is used for protein purification.

c. It serves as a selectable marker for plasmid-containing cells.

how does the polylinker in pUC19 compare to the one in pBR322 in terms of versatility?

a) The polylinker in pUC19 is less versatile

b) The polylinker in pUC19 is equally versatile

c) The polylinker in pUC19 is more versatile

d) Both polylinkers are identical

c) The polylinker in pUC19 is more versatile

What key feature of pUC19 makes it particularly advantageous for cloning experiments?

a) Its large plasmid size

b) Its lack of antibiotic resistance markers

c) Its extensive multiple cloning site (MCS)

d) Its blue-white screening capability

c) Its extensive multiple cloning site (MCS)

What is the purpose of LacZ α-complementation in bacterial plasmids like pUC19?

a) To synthesize β-galactosidase

b) To create blue-colored colonies

c) To allow for blue-white screening of transformed colonies

d) To promote DNA replication

c) To allow for blue-white screening of transformed colonies

Which enzyme's activity is central to LacZ α-complementation screening?

a) DNA polymerase

b) β-galactosidase

c) RNA polymerase

d) Restriction endonuclease

b) β-galactosidase

When bacterial colonies contain the recombinant pUC19 plasmid with a disrupted lacZ gene, what color will they produce on X-gal plates?

a) Blue

b) Green

c) Red

d) White

d) White

Why is LacZ α-complementation important in molecular biology?

a) It allows for the synthesis of colored pigments.

b) It helps identify bacteria species

c) It facilitates the screening of recombinant plasmids with inserted DNA fragments.

d) It promotes DNA replication in bacterial colonies

c) It facilitates the screening of recombinant plasmids with inserted DNA fragments.

In blue-white screening of bacterial colonies using X-gal and IPTG, what is the purpose of X-gal?

a) To promote the growth of bacterial colonies

b) To act as a substrate for β-galactosidase

c) To inhibit the growth of bacterial colonies

d) To serve as a molecular mimic of allolactose

b) To act as a substrate for β-galactosidase

Why do colonies containing the intact pUC19 plasmid (without an inserted DNA fragment) produce blue colonies on X-gal plates?
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a) They have a disrupted lacZ gene.

b) They cannot produce β-galactosidase.

c) They can produce functional β-galactosidase.

d) They lack IPTG.

c) They can produce functional β-galactosidase.

What is the role of IPTG in blue-white screening using X-gal?

a) To inhibit the growth of bacterial colonies

b) To serve as a substrate for β-galactosidase

c) To disrupt the lacZ gene

d) To act as a molecular mimic of allolactose and induce lac operon expression

d) To act as a molecular mimic of allolactose and induce lac operon expression

Which of the following is NOT a purpose of blue-white screening in molecular biology research?
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a) Identifying bacterial colonies containing recombinant plasmids

b) Detecting the presence of functional lacZ genes

c) Confirming successful DNA transformations

d) Synthesizing chromogenic substrates

d) Synthesizing chromogenic substrates

which plasmid would be more suitable for cloning larger DNA fragments?
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a) PBR322

b) pUC19

c) Both are equally suitable for cloning larger fragments.

d) Neither is suitable for cloning larger fragments.

a) PBR322