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Molecular Biology > Hybridization Techniques > Flashcards

Hybridization Techniques Flashcards

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1
Q

It is the phenomenon in which DNA/RNA/Protein anneal to complementary DNA/RNA/Protein

A

Hybridization techniques

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2
Q

It recognizes specific base sequences and cuts the DNA. It is also considered an endonuclease that is naturally found in bacterial cells.

A

Restriction Enzymes

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3
Q

What are the 4 Classifications of Restriction Enzymes?

A

Type I - IV

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4
Q

It contains both nuclease and methylase activity and can recognize specific sequences and cut DNA at a NON-SPECIFIC SITE > LESS THAN 1000 BP AWAY

A

Type I

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5
Q

It is commonly used in the laboratory and is considered unfunctional that can recognize (blank) and cut within the palindrome

A

Type II; Palindromic sequence

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6
Q

It is bifunctional and can recognize specific 5-7 bp sequences and cut 24-24 bp down

A

Type III

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7
Q

What are the different blotting techniques?

A

SNoW
Southern Blots
Nothern Blots
Western Blots

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8
Q

Macroarray and Microarray are types of?

A

Genomic Array Technology

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9
Q

It is the binding of complementary sequences and the formation of a duplex between 2 complementary nucleotide sequences

A

Hybridization

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10
Q

Probes are used for?

A

Detection of specific sequence

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11
Q

What is a primer?

A

It is the starting point of Replication (RNA) ; in vitro = Amplification (DNA)

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12
Q

It is the process of transferring sample nucleic acids on a solid support and is widely used for the identification of desired DNA, RNA fragments, and proteins from thousands of molecules.

A

Blotting techniques

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13
Q

Southern blot detects?

A

DNA

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14
Q

(blank) is isolated and cut with restriction enzymes

A

DNA

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15
Q

DNA fragments are separated by?

A

Gel electrophoresis

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16
Q

It is the process of removing purine bases that has Larger DNA fragments (>500 bp)

A

Depurination

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17
Q

The gel is treated with (blank) on Denaturation

A

NaOH

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18
Q

To move the DNA from the gel to a membrane substrate from probing

A

Blotting (Transfer)

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19
Q

Give 2 membrane types

A

Pure nitrocellulose and Reinforced nitrocellulose

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20
Q

Give 3 transfer methods

A

Capillary transfer, Electrophoretic transfer, and Vacuum transfer

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21
Q

It is driven by the capillary movement of a buffer from the soaked paper to the dry paper.

A

Capillary transfer

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22
Q

driven by the capillary movement of a buffer from the soaked paper to the dry paper, (blank) moves from the gel to the membrane. the (blank) will adhere to the membrane, which will be subsequently exposed to the probe.

A

denatured DNA; DNA

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23
Q

This system uses electric current to move the DNA transversely through the get to the membrane. This type of transfer is used mostly for small fragments or proteins.

A

Electrophoretic System/Transfer

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24
Q

This system uses suction and buffer recirculation to move the DNA out of the gel and onto the membrane.

A

Vacuum transfer

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25
Q

It is generally faster than capillary transfer for (blank) DNA fragments. However, unlike capillary transfer, (blank) requires specialized equipment.

A

large; vacuum transfer

26
Q

A process to prevent the transferred DNA fragments from washing away or moving on the membrane

A

Immobilization of DNA

27
Q

Bake the membrane in a (blank) oven at what degrees with time?

A

Vacuum; 80*C - 30 to 60 mins

28
Q

This prevents the probe from binding to nonspecific sites on the membrane surface

A

Prehybridization

29
Q

True or False:

Buffer does contain probe in Prehybridization

A

FALSE

Buffer does NOT contain probe in Prehybridization

30
Q

True or False:

Before hybridization, the excess probe is washed from the membrane

A

FALSE

AFTER hybridization, the excess probe is washed from the membrane

31
Q

It is when the pattern of hybridization is visualized

A

Detection

32
Q

What are the two ways to detect patterns of hybridization?

A

Xray film and Chromogenic detection method

33
Q

It is the modification of the Southern blot technique

A

Northern blots

34
Q

This blotting technique is designed and developed by James Alwine to investigate RNA

A

Northern blots

35
Q

This blotting technique is another modification of the Souther blot that immobilized proteins

A

Western Blots

36
Q

What is the target cell of Southern blot and its probe?

A

DNA ; Nucleic Acid

37
Q

What is the target cell of Northern blot and its probe?

A

RNA ; Nucleic Acid

38
Q

What is the target cell of Western blot and its probe?

A

Protein ; Protein

39
Q

What is the target cell of Southwestern blot and its probe?

A

Protein ; DNA

40
Q

What is the purpose of Southern blot?
A. Gene Structure
B. Protein processing, gene expression
C. Transcript structure, processing, gene expression
D. DNA binding proteins, gene regulation

A

A. Gene Structure

41
Q

What is the purpose of Northern Blot?
A. Gene Structure
B. Protein processing, gene expression
C. Transcript structure, processing, gene expression
D. DNA binding proteins, gene regulation

A

C. Transcript structure, processing, gene expression

42
Q

What is the purpose of Western Blot?
A. Gene Structure
B. Protein processing, gene expression
C. Transcript structure, processing, gene expression
D. DNA binding proteins, gene regulation

A

B. Protein processing, gene expression

43
Q

What is the purpose of Southwestern Blot?
A. Gene Structure
B. Protein processing, gene expression
C. Transcript structure, processing, gene expression
D. DNA binding proteins, gene regulation

A

D. DNA binding proteins, gene regulation

44
Q

These are small fragments of nucleic acid that are complementary to a part of the DNA or RNA of the organism we are searching for

A

Probes

45
Q

What are the two types of probe labeling?

A

Radioactively and Fluorescently

46
Q

What are the two types of fluorescent labeling?

A

Biotin and Digoxygenin

47
Q

It is a more specific type of probe that is expensive and difficult to synthesize which may require reaction times of hours to hybridize

A

Long probes (500-5000 bp)

48
Q

It hybridizes at very high rates and is less specific

A

Shorter probes (<500 bp)

49
Q

What are the factors affecting stringency?

A

Temperature, salt concentration of the hybridization buffer, and concentration of denaturant

50
Q

Tm = 4 x number of GC pairs + 2 x number of AT pairs

This formula is used calculate the?

A

Melting temperature

51
Q

The determination of the size of the target is not required and is deposited directly on the membrane

A

Dot/Slot Blot

52
Q

The target is deposited in a circle or dot

A

Dot/Slot Blots

53
Q

The target is deposited in an oblong bar

A

Slot Blots

54
Q

The targets can be screened simultaneously in a very small area by miniaturing the deposition of droplets

A

Microarrays

55
Q

It is used to measure mRNA expression and is more sensitive than Northern blotting. It is also used to detect sequence changes in DNA or mutational analysis.

A

Solution hybridization

56
Q

Fluorescent In Situ Hybridization is also known as?

A

In Situ Hybridization or ISH

57
Q

It is used to detect and localize the presence or absence of specific DNA sequences on chromosomes

A

ISH

58
Q

What are the requirements for FISH?

A

Sample, Fluorescent Probe and Fluorescence Microscope

59
Q

The label is bound to the probe

A

Direct labelling

60
Q

It requires an additional step before detection. The probe was detected using antibodies conjugated to labels.

A

Indirect labeling

61
Q

A fluorescent probe attaches to the target sequence during hybridization and is visualized through a fluorescence microscope

A

Visualization

Molecular Biology (1 decks)

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