dna extraction
DESCRIPTION
Dna Extraction. How does it actually work?. Cell Lysis. Lysis buffers are typically detergents such as sodium dodecyl sulfate (SDS) What happens if you add dish soap to a frying pan with water that has a layer of grease on top?. Lysis Cont’d. Another analogy: popping a water balloon - PowerPoint PPT PresentationTRANSCRIPT
DNA EXTRACTIONHow does it actually work?
Cell Lysis Lysis buffers are
typically detergents such as sodium dodecyl sulfate (SDS)
What happens if you add dish soap to a frying pan with water that has a layer of grease on top?
Lysis Cont’d Another analogy:
popping a water balloon When you pop a
balloon, it is like lysing a cell
When the balloon is popped, all of the contents are released
Lysis Cont’d So what actually
happens? Detergents disrupt
lipid-lipid interactions compromising cell membrane integrity
Membrane is destroyed or becomes “leaky” depending on lysis buffer
Wash Stage The next stage typically involves
solutions to remove excess salts and impurities
Additionally, proteases, Rnases, etc. are often added to remove proteins (e.g., histones), RNA, etc.
Wash Cont’d The wash solution (though separate
solutions are sometimes used before) also adjusts pH
At low pH, DNA binds to a column Historically made of silica—a combination of
salts, silica and low pH create conditions that attract DNA
Elution Elution is a fancy
term for release of DNA from column
Water or TE (or other elution buffer) has a neutral or alkaline pH, leading to release of DNA
QIAGEN Kit A few differences:
Column properties differ: lysing action
Column proprietary; not the traditional silica column