Practical 2 Flashcards
how much of human DNA codes for actively produced proteins?
1%
what ploidy are bananas and features:
triploid 3n, three sets of chromosomes
- infertile (unequal no.)
general experiment flow:
- mush bananas
- add extraction buffer
- add meat tenderiser
- filter solution into falcon tube using cheesecloth filter funnel
- add ethanol onto side
- two layers will form
- take out DNA into ethanol in Eppendorf tube
what was the extraction buffer made of:
detergent and salt
purpose of detergent in extraction buffer:
solubises cell membrane, break down lipids of cell membrane
purpose of meat tenderiser:
protease, break down proteins
purpose of ethanol:
make DNA insoluble, precipitates it for easy extraction
REAGE stands for:
restriction endonucleases and agarose gel electrophoresis
what are restriction enzymes:
or restriction endonucleases
- reduces the size of DNA molecule, for easier analysis
- type of enzyme good for this
what are restriction enzymes good for:
act as molecular scissors, cut at restriction sites (specific nucleotide sequences)
- 4-6 bp long
- restriction sites are palindromes
specific application of restriction enzymes:
restriction fragment length polymorphism (RFLP)
how does electrophoresis work:
charged particles will move across a medium in a electrical field
- rate of movement will depend on size, charge etc.
- need gel like agarose as support medium
how will the substance be separated in the agarose gel matrix:
- nucleic acids sorted based on size
how was the pathogenic and non pathogenic bacteria E coli different?
- seen in the DNA bands of the RFLP image non pathogenic (N) had more and smaller fragments while (P) had less restriction sites, thus less bands due to mutations
electrophoresis eg.
- DNA has negative charge (from sugar phosphate backbone), will migrate to cathode (positive charge)
- longer fragment will move slower and get caught in gel matrix
