Modification of standard proteinase K/phenol method for extraction of DNA from small tumour biopsies.
The standard proteinase K/phenol DNA isolation method was found to produce unsatisfactory yields of DNA from small tissue biopsies (less than 50 mg). The influences of the volume of cell lysis buffer and the amount of proteinase K on the final DNA yield and quality were studied, and an improved method was devised and compared with both the standard procedure and a phenol-free protocol. The optimal volume of cell lysis buffer was found to be 200 microliters per mg of tissue while the optimal amount of proteinase K was 60 micrograms per mg of tissue. A mean yield of 12 mu/mg tissue of pure, high molecular weight DNA was achieved from 50 frozen samples prepared by crushing. Yields from 20 microns thick cryostat sections reached 30 micrograms/mg.
Pitera
R
R
Department of Histopathology, King's College School of Medicine and Dentistry, Denmark Hill, London, UK.
Pitera
J E
JE
Mufti
G J
GJ
Salisbury
J R
JR
D000433:1-Propanol; D001706:Biopsy; D004273:DNA, Neoplasm / Q000032:analysis*; D019286:Endopeptidase K; D006801:Humans; D008228:Lymphoma, Non-Hodgkin / Q000737:chemistry / Q000235:genetics* / Q000473:pathology; D008722:Methods; D010636:Phenols; D012189:Retrospective Studies; D012697:Serine Endopeptidases; D013160:Splenic Neoplasms / Q000737:chemistry / Q000235:genetics* / Q000473:pathology; D013274:Stomach Neoplasms / Q000737:chemistry / Q000235:genetics* / Q000473:pathology
D004273:DNA, Neoplasm; D010636:Phenols; D000433:1-Propanol; D012697:Serine Endopeptidases; D019286:Endopeptidase K