Gene Technology Flashcards
What is a Palindromic site?
its read the same forwards and backwards
eg) CTTAAG to GAATTC
What is the role of restriction enzymes?
They cut DNA at specific Palindromic sites called restriction sites
What does using a restriction enzyme leave DNA with?
sticky ends (unpaired bases)
What does reverse transcriptase do?
mRNA ——-> cDNA
What does cDNA not have?
introns (made from mRNA post splicing)
What enzyme does DNA ——> mRNA?
RNA Polymerase
Describe Artificial Synthesis of a gene.
use a gene machine to make DNA from scratch, making oligonucleotides, join theses together to make a synthetic gene.
What is an oligonucleotide?
a sequence of around 25 nucleotides
What are the three ways we can isolate target genes?
gene machine, reverse transcriptase or restriction enzymes
What is PCR used for?
to amplify a gene
What are the 4 components of PCR?
DNA sample, free nucleotides, primers, DNA polymerase
describe the 3 steps of PCR
heat up to 95 degrees
cool to 50 degrees
heat to 70 degrees
Why do we heat the sample to 95 degrees
break the H+ bonds between so they are single stranded
Why do we cool the sample to 50 degrees
allows the primer to attach by complementary base pairing to the exposed strand - making a double strand - so DNA polymerase can bind to the double stranded section
Why do we heat the sample back up to 70 degrees
there is a higher kinetic energy increasing enzyme action
why can the DNA polymerase work at high temperatures?
extracted from thermophillic bacteria
What is a primer?
Short sequence of DNA which is complementary to the target DNA
Name three ways we can isolate a target gene
restriction enzymes, Gene machine, reverse transcriptase
What charge is DNA?
negative
Describe the process of electrophoresis
attach a fluorescent label to DNA
put labelled DNA fragments into the well at the negative end
turn on current
DNA is attracted to the positive electrode
How can we distinguish the length of DNA from electrophoresis?
smaller fragments move faster (therefore further over any given time)