Recombinant DNA 1 Flashcards
What is genomic DNA?
Chromosomal DNA that is long, contains introns and used for genome mapping and sequencing
How is cDNA synthesised?
From mRNA in a tube using reverse transcriptase
What does the synthesis of cDNA require?
A primer and the nucleotides (dATP, dGTP, dCTP and dTTP)
What is plasmid DNA?
Bacterial extrachromosomal DNA, circular and used as a vector
How are restriction enzymes/ endonuclease made?
From bacteria
What is the role of restriction enzymes/ endonuclease?
Cut double stranded DNA at defined sites
What sequences do restriction enzymes/ endonuclease produce?
Palindromic
How long are recognition sequences?
4, 6 or 8 nucleotides
What are restriction enzymes named after?
The species of bacteria which it was isolated from
What make it easier for hydrogen bonds to from with external DNA?
Sticky ends
What are the stages of using a restriction endonuclease?
DNA from two sources is isolated and cleaved with the same restriction endonuclease
Two types of DNA pair at their sticky ends. DNA ligase joins the segments.
The plasmid is inserted into a bacteria cell by transformation; cells are grown on a plate
Clones are screened for genetic interest.
What is the role of DNA ligase?
Joins DNA fragments by ligating the phosphodiester backbone
What does blue white screening identify?
Antibiotic resistance cells which also contain recombinant DNA
What does the enzyme ß-galactosidase (lacZ) convert?
Substrate X-gal into a blue-colored product
