A protocol for  high-quality genomic DNA extraction from legumes.
Current DNA extraction protocols, which require liquid nitrogen, lyophilization and considerable infrastructure in terms of instrumentation, often impede the application of biotechnological tools in less researched crops in laboratories in developing countries. We modified and optimized the existing CTAB method for plant genomic DNA extraction by avoiding liquid nitrogen usage and lyophilization. DNA was extracted directly from freshly harvested leaves ground in pre-heated CTAB buffer. Chloroform:isoamyl alcohol (24:1) and RNase treatments followed by single-purification step decontaminated the samples thereby paving way for selective extraction of DNA. High molecular weight DNA yield in the range of 328 to 4776 ng/μL with an average of 1459 ng/μL was obtained from 45 samples of cultivated and wild Cajanus species. With an absorbance ratio at 260 to 280 nm, a range of 1.66 to 2.20, and a mean of 1.85, very low levels of protein and polysaccharide contamination were recorded. Forty samples can be extracted daily at a cost between 1.8 and US$2.0 per plant sample. This modified method is suitable for most plants especially members of the Leguminosae. Apart from Cajanus, it has been extensively applied in DNA extraction from Cicer and Vigna species.
Crop Improvement Division, Indian Institute of Pulses Research, Kanpur, India. ikechukwu.agbagwa@gmail.com
I O
IO
Agbagwa
S
S
Datta
P G
PG
Patil
P
P
Singh
N
N
Nadarajan
D002021:Buffers; D000077286:Cetrimonium; D002593:Cetrimonium Compounds / Q000737:chemistry; D002725:Chloroform / Q000737:chemistry; D018744:DNA, Plant / Q000235:genetics / Q000302:isolation & purification*; D007887:Fabaceae / Q000235:genetics*; D018745:Genome, Plant; D018895:Microsatellite Repeats; D000439:Pentanols / Q000737:chemistry; D018515:Plant Leaves / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D052616:Solid Phase Extraction; D012997:Solvents / Q000737:chemistry
D002021:Buffers; D002593:Cetrimonium Compounds; D018744:DNA, Plant; D000439:Pentanols; D012997:Solvents; D002725:Chloroform; C029683:isopentyl alcohol; D000077286:Cetrimonium