Cloning of DNA L5

Cloning and DNA flash cards.
Word on one side
Description/answer to question on other 
A Level and University level 

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  • Created by: Alex
  • Created on: 11-04-13 15:11
Host Cell
Cell that is carrying around foreign DNA or RNA
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Complementary DNA (cDNA)
DNA synthesized from mRNA via reverse transcriptase and DNA polymerase
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Reverse transcriptase
is an enzyme used to generate complementary DNA (cDNA) from an RNA template, a process termed reverse transcription
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DNA polymerase
enzyme that catalyzes the polymerization of deoxyribonucleotides into a DNA strand. DNA polymerases are best-known for their role in DNA replication, in which the polymerase "reads" an intact DNA strand as a template and uses it to synthesize the new
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dsDNA
Double stranded DNA
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ssDNA
Single Stranded DNA
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Selectable marker
a gene introduced into a cell that confers a trait for artificial selection
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DNA cloning
Selectively amplifying DNA sequences to their structure and function can be studied
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Method of DNA cloning
Inserting foreign DNA into vectors capable of replicating in host cell (e.g. E.coli) - host cell grows and divides producing multiple clopies of identical DNA
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How to insert DNA in cloning
Restriction enzymes, T-A , TOPO, Recombination based
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Restriction enzymes in DNA cloning
DNA cut with blunt or sticky end restriction enzymes. Plasmind cut with same enzyme. - complementary cut ends. DNA ligase joins DNA into plasmid DNA
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T-A DNA cloning
relies on adenine and thymine (complementary basepairs) on different DNA fragments to hybridize and, become ligated together. PCR creates insert and adds AAA overhang. vector cut with blunt end ddTTP leaves TTT overhang. Faster . no choice directio
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Requirements for DNA cloninh
Foreign DNA. Host organism. Vector DNA for cloning. Way of inserting DNA in/out of vectore. way of placing modified DNA into host cell. Methods for selecting cells with foreign DNA
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Vector DNA
DNA molecule that functions as molecular carrier - brings DNA of interest into host cell.
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Competent (cells)
is the ability of a cell to take up extracellular ("naked") DNA from its environment
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Transfering DNA into the host cell
Tansformation, Electroporating, Conjugation, Transfection
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Transformation
Cells are heatshocked forcing the DNA to enter through cell pores or damaged memberanes
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Electroporating DNA into host cell
Membranes permeabilized by electric field - means DNA can enter
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Conjugation DNA into host cell
natural transmission from donor to recipient - cell to cell junctions formed
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Transfection
DNA is packed in vitro into phage particles. phages infect bacterial cells and DNA is transiently expressed
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Selecting and screening recombinants
identified by selective growth conditions
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Different selective growth conditions.
Antibiotics, nutrient requirements, plaque type, Blue-White selection (Beta-galactosidase)
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Things needed to reprogramme a cell
Reactivate suppressed ganes and activate other genes
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Other cards in this set

Card 2

Front

Complementary DNA (cDNA)

Back

DNA synthesized from mRNA via reverse transcriptase and DNA polymerase

Card 3

Front

Reverse transcriptase

Back

Preview of the front of card 3

Card 4

Front

DNA polymerase

Back

Preview of the front of card 4

Card 5

Front

dsDNA

Back

Preview of the front of card 5
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