Flashcards in Biotechnology - PCR and Gel electrophoresis Deck (10)
PCR or polymerase chain reaction a technique used in molecular biology for producing multiple copies of DNA from a sample; used in DNA fingerprinting and in identifying diseases
Process of PCR
- PCR uses the heat to separate the DNA into two strands allowing each to be copied. its is completed at 94-97 degrees to separate the strands without disrupting each individual strand.
- The temperature drops to 50-60 degrees which allows for short strands of DNA called primers to bind to the single DNA strands.
3. Extension or Elongation;
- DNA polymerase is used to join new, complementary nucleotides to the sections origination with the primers. This extends to nucleotide chain and creates a new strand of DNA however, it is not full length as its starts from the primer. Taq polymerase is used as it does not denature at hot temperatures which allows the process to be simplified and automated.
A technique that is able to separate DNA strands based on their lengths. The DNA are placed in Wells (an indentation in the gel) in a semi-solid gel that is immersed in a solution of an electrolyte. Negative electrode is closest to the DNA and positive is furthers away. So when the current is passed through the Negatively charged DNA moves to the positively charged DNA. Smaller strands move faster than longer DNA strands.
When added to the DNA, they cut the strands into different lengths depending on their base sequence
The binding pattern of the DNA strands. DNA profiling is a technique that uses binding patterns of DNA fragments as a means of identification; a DNA fingerprint is a unique to a particular individual also known as a DNA profile
A 'run' at the same time as the samples. Contains segments of DNA with known lengths and is compared with the results of unknown samples to determine the lengths of the DNA strands in the sample
is the determination of the precise order of nucleotides in a sample of DNA.
What happens when a DNA forms.
When DNA forms each nucleotide loses two phosphate groups. The Sugar molecule also loses a Hydrogen atom from the hydroxy group (OH) when it bonds to the phosphate group of an adjacent nucleotide.
Sangers method of DNA sequencing
nucleotides that lack the OH are added to the growing strand. These are called dideoxy-ribonucleotides. They stop elongation of the sequence because there is no OH group for the next nucleotide to attach on to. This creates varying lengths of DNA which can be separated through gel electrophoresis