DNA Purification Flashcards
(16 cards)
what are the principles of dna extraction
sample source
lysis
purification
concentration
what are the two methods of cell lysis and what are some examples
chemical - solvents, enzymes and detergents
mechanical - sonification, beads
what is the organic solvent method of purification
using phenol to denature proteins and chloroform to cause phase separation
what are some enzymatic forms of purification
using dnase or rnase, as well as proteinase k
what is the sillica bead version of purification
using sillica beads to attach the dna, strong detergent alters debris but not DNA, washed, then DNA removed using elution buffer
what can be used to make dna precipiotate out of solution
ethanol or isopropanol and salt
what is the spool method of concentration
using a glass spool to pull long DNA molecules out of solution
how can you isolate plasmid dna
by nicking larger supercoiled dna and centrifuging, leaving plasmid DNA in supernatent
what are the differences in terms of durability of plasmid and chromosomal dna
plasmid dna is more resisrant to nicking than chromosomal dna
what is the acid base purification of plasmids
using sodium hydroxide, then acid, linear dna is coagulated then separated by centrifugation
how can phage dna be purified
using polyethylene glycol and salt
what can be used to measure dna purity
spectrophotometry
what are the wavelegnths used to assay DNA ourity
260 and 280
what are some differences between dna and rna
rna is shorter and more easily degraded, with enzymes such as rnase
what is the threshold for purity in wv rationbs
1.8 or higher is pure
what can be used to separate mrna
in eukaryotes, poly a tails can be used, or oligio dts
