topic 8 recombinant DNA technology Flashcards
what is recombinant DNA technology?
when a fragment of DNA is transferred from one organism to another
why can fragments of DNA be transferred between organisms?
genetic code is universal so the same DNA base triplets code for the same amino acids in all living organisms
transcription and translation mechanisms are similar
what are the three ways in which DNA fragments can be made?
using reverse transcriptase
using restriction endonuclease enzymes
using a gene machine
how can reverse transcriptase be used to make DNA fragments?
cells only have 2 copies of each gene but contain many mRNA molecules complementary to the gene so easier to obtain
mRNA molecules can be used as templates to make lots of DNA
reverse transcriptase enzymes make DNA from RNA template
DNA produced is complementary (cDNA)
mRNA isolated from cells, mixed with free DNA nucleotides and reverse transcriptase, which uses mRNA as a template to synthesise a new strand of cDNA
how can restriction endonuclease enzymes to make DNA fragments?
some sections of DNA have palindromic sequences of nucleotides
sequences consist of antiparallel base pairs
restriction endonucleases are enzymes that recognise specific palindromic sequences (recognition sequences) that cut DNA at these places
different restriction endonucleases cut at different specific recognition sequences, as the shape of the recognition sequence is complementary to the enzyme’s active site
if the recognition sequences are present at either side of the DNA fragment, you can use restriction endonucleases to separate the rest from DNA
DNA sample incibated with specific restriction endonuclease- cuts DNA fragment out by a hydrolysis reaction
sometimes leaves sticky ends
what are sticky ends?
small tails of unpaired bases at the end of the fragment- can bind the DNA fragment to another piece of DNA that has sticky ends with complementary sequences
how can a gene machine be used to make DNA fragments?
technology has been developed so fragments of DNA can be synthesised from scratch, without needing a pre-existing DNA template
database contains the necessary information to produce the DNA fragment
any sequence can be made
what are the stages of a gene machine?
sequence required designed
first nucleotide in the sequence is fixed
nucleotides are added step by step in the correct order in a cycle of processes
eg) add protecting groups, ensures that nucleotides are joined at the right points to prevent unwanted branching
what are olgionucleotides?
short sections of DNA
around 20 nucleotides long
once complete broken off from the support and protecting groups are removed
can be joined to make longer DNA fragments
