In Vitro Gene Cloning Flashcards
What is the PCR a method of?
Copying fragments of DNA
What method does in vitro cloning use?
The polymerase chain reaction
What does the PCR require in order to take place?
The DNA fragment to be copied, DNA polymerase, primers, free nucleotides and a thermocycler which is a computer-controller machine that varies temperatures precisely over a period of time
What is the first stage of the PCR?
The DNA fragments, primers and DNA polymerase are placed in a vessel in the thermocycler which heats the mixture to 95 degrees - separating the two strands of the DNA fragment
What is the second stage of the PCR?
The mixture is cooled to 55 degrees causing the primers to join (anneal) to their complementary bases at the end of the DNA fragment
What is the importance of primers?
They provide the starting sequences for DNA polymerase to begin DNA copying because DNA polymerase can only attach nucleotides to the end of an existing chain and they also prevent the two separate strands from simply rejoining
What is the third stage of the PCR?
The temperature is increased to 72 degrees which is the optimum temperature for the DNA polymerase to add complementary nucleotides along each of the separated DNA strands
How long does one heat cycle of the PCR take?
2 minutes
What are some of the advantages of in vitro cloning?
It is very fast, does not require living cells and it has very useful applications in forensics
What are some of the advantages of using in vivo cloning?
It is very useful when we wish to introduce a gene into another organism, It involves almost no risk of contamination, it is very accurate, it cuts out specific genes and produces transformed bacteria that can be used to produce large quantities of gene products
