Simultaneous extraction of nuclear and mitochondrial DNA from human blood.
A method of simultaneous isolation of nuclear DNA (nDNA) and mitochondrial DNA (mtDNA) from human blood has been proposed by improvising Lahiri's method of isolation of nuclear DNA. The approach presented here provides selectively enriched fractions and eliminates the need for two different methods or separate reagent sets for the extraction of nDNA and mtDNA. It employs an initial nuclear/ cytoplasm partitioning, followed by the similar procedural steps for the two fractions separately. It gives good quality and quantity of the nDNA as well as the mtDNA, suitable for processes like PCR amplification and sequencing and may prove to be useful for people studying population genetics and evolution using molecular markers maximizing the available resources, especially in cases where a large database needs to be generated from limited amount of blood sample. From 3 ml of blood, the yields of mtDNA salvaged from the supernatant were sufficient to set approximately 4x10(5) reactions (starting with 250 fg DNA per reactions) of mtDNA loci which otherwise would have been discarded as per original Lahiri's procedure. The quality of mtDNA from the mitochondrial fraction was suitable for all major downstream processes as confirmed by locus specific PCR amplifications and sequencing. Through this procedure, the wastage of nDNA can be avoided when mtDNA loci is studied.
Ahmad
Shazia
S
Low Level Radiation Studies Section, Radiation Biology and Health Science Division, Bio-Medical Group, Bhabha Atomic Research Center, Mumbai, India. shaziaahmad_12@yahoo.co.in
Ghosh
Anu
A
Nair
Divya L
DL
Seshadri
Murlidhardas
M
D002467:Cell Nucleus / Q000737:chemistry; D004247:DNA / Q000097:blood* / Q000302:isolation & purification*; D004272:DNA, Mitochondrial / Q000097:blood* / Q000302:isolation & purification*; D006801:Humans; D016133:Polymerase Chain Reaction
D004272:DNA, Mitochondrial; D004247:DNA