DNA Extraction Notes

Did small test using supernatant from centrifugation step with K 1 (in Koba Extraction) samples #62, 63 (Sheet BH)
Results: Got DNA pellet.
Need to check to see what yield looks like and how it works for PCR.

Prep, pptn etc

Prepared leaf tissue (BG 1-36) for DNA extraction tomorrow.
Autoclaved tubes, tips for extraction.

Precipitated DNA for extracts BF 37-72: Samples BF 47, 55 and 71 had very tiny pellets prior to centrifugation.

DNA Extraction etc

Sheet BF 37-72 last fp#: 3336
Leaf tissue for extraction prepared on previous day.

Changed waterbath water
Gave MB list of items to order.
Prepared tips & tubes for autoclaving.

Questions

How long does 0.5M EDTA (pH 8.0) last on benchtop/in fridge?
Other chemicals whose expiration/half-life is unknown: Sorbitol (solid and 1.75M prep), CTAB, N-Lauroyl Sarcosine.

About

A CRU DNA Lab chronicle of observations, notes, protocols, recipes etc and other general information which maybe useful to archive.

DNA Extractions

Chemicals used (approximation)

70 mL CIAA per 100 extracts

100 mL Isopropanol per 100 extracts

100 mL Ethanol per 100 extracts


Notes

D-squared note: Overnight pptn of DNA ok. Pellets unaffected.

EDTA Used in March 2002: Disodium dihydrate

Gel quantitation: Dryloading works well only if yield is good.

PCR and Electrophoresis

PCR Notes

TAQ + Buffer + Magnesium Chloride 50U/mL: 0.1 mL per reaction

Group A: 27/28; 45A/45B; 53/54B; 132/133
Group B?: 240/241; 246/247
Group C: 86/87
Group D: 35/36; 43/44

Electrophoresis Notes

Ladder (Lambda) 114ng/mL: 25 mL per gel

Long Ranger gels
Sigma Urea dissolves in