In Vivo/In Vitro Cloning A2 Flashcards
how to isolate specific genes from human DNA
use restriction endonuclease
to cut DNA at specific base sequence
define recombinant DNA
contains genes from two species or types of organism
what does reverse transcriptase do
makes cDNA from RNA
what is restriction endonuclease
enzyme that cuts DNA into smaller fragments
cuts at specific base sequence
breaks phosphodiester bonds
what is the purpose of sticky ends
to join 2 pieces of DNA by complementary base-pairing/binding
process of making recombinant DNA
cut desired gene from another organism
using restriction endonuclease to make sticky ends
use the same enzyme to cut plasmid
mix together and use DNA ligase to join wanted gene to plasmid
how to put a plasmid in a bacterial cell
place in solution of calcium ions to increase permeability of membrane, allowing plasmid to be taken up
how to identify bacteria with recombinant DNA
plasmids contain gene for fluorescence
plasmids contain antibiotic resistant gene which acts as marker gene
why is it good to use a fluorescent marker gene
allows quick identification of transformed bacteria under UV light
define vector
carrier of DNA into another organism
example of a vector
plasmid
what is a DNA probe
single stranded DNA
base sequence complementary to DNA that is being identified
radioactively/fluorescently labelled so it can be detected
define in vitro cloning
polymerase chain reaction
used to make many copies of gene or amplify small amount of DNA
temperature changes in polymerase chain reaction
95
52
72
why is PCR first heated to 95 degrees
H bonds break between double stranded so polynucleotides strands separate
