RECOMBINANT DNA TECHNOLOGY

Question 1

TECHNOLOGY THAT ENCOMPASSES A NUMBER O F EXPERIMENTAL PROTOCOLS
LEADING T O TRANSFER O F DNA FROM ONE ORGANISM T O ANOTHER

Answer 1

Recombinant DNA TECHNOLOGY

Question 2

Molecular scissors that cut DNA into fragments at specific sites in their sequence.

Answer 2

Restriction endonucleases

Question 3

EcoRI
“Eco” word from?

Answer 3

1st letter of genus +
1st and 2nd letter
of species

Question 4

EcoRI
“R”from ?

Answer 4

Strain of host bacteria

Question 5

EcoRI
“I”from ?

Answer 5

Order of discovery

Question 10

CLASSIFICATION OF RESTRICTION ENZYMES

Cut DNA at random far from their recognition sequences.

Answer 10

Type 1

Question 10

Cut DNA at defined positions close to or within their recognition sequences.

Answer 10

Type 2

Question 10

Cut modified (e.g. methylated) DNA.

Answer 10

Type 4

Question 11

Cut DNA outside of two recognition sequences in opposite orientations.

Answer 11

Type 3

Question 12

MECHANISM OF RESTRICTION ENZYMES
1.
2.
3.

Answer 12

1. SCAN - Look fo a particular pattern of
   nucleotides:ENZYME’S RECOGNITION
   SEQUENCE
2. BIND- The enzyme will bind the DNA
   molecule.
3. CUT- The enzyme cuts in each of the two
   sugarphosphate backbones of the
   double helix

Question 13

Alul and Haelll produce _____ ends.

Answer 13

blunt

Question 14

DNA sequence that produce blunt ends.

Answer 14

Alul and Haelll

Question 15

BamHl, Hindlll and EcoRi produce ____ ends

Answer 15

sticky

Question 16

Made by fusing DNA-binding domain to a nuclease domain.
Ex. CRISPR technolog

Answer 16

Artificial Restriction enzymes

Question 16

DNA sequence that produce “ sticky” ends

Answer 16

BamHl, Hindlll, and EcoRi

Question 17

Who won nobel prize in Chem 2020, “for the development of a method for genome editing.

Answer 17

Emmanuelle Charpentier
Jennifer A. Doudna

Question 18

A DNA MOLECULE IN WHICH FOREIGN DNA
MOLECULE IS INSERTED AND WHICH IS FURTHER CAPABLE OF REPLICATING WITHIN HOST CELL TO PRODUCE MULTIPLE CLONES OF THE RECOMBINANT DNA.

Answer 18

CLONING VECTOR

Question 19

MAIN FEATURES OF CLONING VECTORS

Answer 19

1. ORIGIN OF REPLICATION
2. MULTIPLE CLONING SITES
3. SELECTABLE MARKER

Question 20

DNA segment with several RE sites located next to each other.

Answer 20

MULTIPLE CLONING SITE

Question 21

Allows selection of host cells with the recombinant vector.

Answer 21

Selectable marker

Question 22

CLONING STRATEGIES _________ involves the ligation of short, staggered ends that can be rejoined by DNA ligase to regenerate original recognition sequence.

Answer 22

Cohesive End Cloning

Question 23

CLONING STRATEGIES _____ involves ligating dsDNA into a plasmid where both the insert and linearized plasmid have no overhanging bases at their termini.

Answer 23

Blunt End Cloning

Question 24

10-100x less efficient than cohesive-end cloning. Desired insert does not require any restriction sites.

Answer 24

Blunt End Cloning

Question 25

CLONING STRATEGIES Modified Blunt End Cloning may use…

Answer 25

* Linkers * Adaptors * Homopolymer tails

Question 26

final step in the construction of a recombinant DNA molecule where the vector molecule and the DNA to be cloned are joined together by DNA ligase.

Answer 26

LIGATION

Question 27

Two families of dna ligases

Answer 27

1.ATP-dependent Ligase 2. NAD+-dependent ligases (LigA)

Question 28

All known eukaryal cellular DNA ligases; also in archaea.

Answer 28

ATP-dependent Ligase

Question 29

distinctive and structurally homogeneous clade of enzymes found in all bacteria.

Answer 29

NAD+-dependent ligases (LigA)

Question 30

is the process by which an organism (competent) acquires exogenous DNA.

Answer 30

TRANSFORMATION

Question 31

It is a Direct Methods of transformation, * CaCl2 * TSS Buffer with PEG, DMSO, and MgCl2 * Usually followed by heat-shock transformation.

Answer 31

Chemical Competency

Question 32

It is a Direct Methods of transformation, _____ significantly increases the electrical conductivity of cell membrane for subsequent increase in its permeability.

Answer 32

Electrocompetency

Question 33

It is a Direct Methods of transformation, the use of a glass micropipette to inject a liquid substance at a microscopic or borderline macroscopic level.

Answer 33

Microinjection

Question 34

It is a Direct Methods of transformation, * Transforming yeast cells by limited physical disruption in the presence of DNA * This method is less efficient but considerably more convenient

Answer 34

Agitation by glass beads

Question 35

4 INDirect Methods of transformation.

Answer 35

Transduction Agrobacterium Mediated Protoplast Fusion Conjugation

Question 36

2 METHODS OF SELECTION

Answer 36

1.Use of markers 2.Use of reporter genes

Question 37

ANTIBIOTICS -Positive selection * Ex. Ampicillin under what methods if selection?

Answer 37

METHODS OF SELECTION: 1. USE OF MARKERS

Question 38

Negative selection

Answer 38

BLUE-WHITE SCREENING

Question 39

The genes encode protein products that can be observed using certain biochemical assays.

Answer 39

METHODS OF SELECTION: 2. Reporter genes

Question 40

Examples of Reporter genes. The genes encode protein products that can be observed using certain biochemical assays.

Answer 40

– β-glucuronidase (GUS) – Luciferase gene – Green Fluorescence Protein (GFP)