DNA Bank

Boyd Orr

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DNA bank (n = 734)

DNA was extracted from 4 ml K-EDTA venous blood and quantitated by picoGreen assay, validated by optical density 260nm/280nm checks, and concentrations were equalized. Long-term stock DNA aliquots were laid down, and working 96-well plates of DNA dilutions to 10 ng/µl were prepared. These dilutions are suitable for direct usage, for example 5ng PCR from genomic DNA, for preparation of long PCR sub-banks of specific gene regions and for genomewide preamplifications (using degenerate oligo primer methods or phi29 methods) to conserve stock DNA.

References:

Miller SA, Dykes DD, Polesky HF. A simple salting out procedure for extracting DNA from human nucleated cells Nucleic Acids Res. 1988;16(3):1215.

Cheung VG, Nelson SF. Whole genome amplification using a degenerate oligonucleotide primer allows hundreds of genotypes to be performed on less than one nanogram of genomic DNA. Proc Natl Acad Sci 1996;93(25):14676-9.

Gu DF, Hinks LJ, Morton NE, Day IN. The use of long PCR to confirm three common alleles at the CYP2A6 locus and the relationship between genotype and smoking habit. Ann Hum Genet. 2000;64(Pt 5):383-90.

Dean FB, Nelson JR, Giesler TL, Lasken RS. Rapid amplification of plasmid and phage DNA using Phi 29 DNA polymerase and multiply-primed rolling circle amplification. Genome Res. 2001;11(6):1095-9.

Ahn SJ, Costa J, Emanuel JR. PicoGreen quantitation of DNA: effective evaluation of samples pre- or post-PCR.Nucleic Acids Res. 1996;24(13):2623-5.

Day, I.N.M. (editor) “Molecular Genetic Epidemiology: A Laboratory Perspective” Cambridge, Springer-Verlag, Berlin, Heidelberg, New York 2001.