Molecular characterization of bacterial leaf blight resistant
Table 3: Jaccard’s similarity coefficient of 30 rice genotypes based on RAPD data analysis
(TE) buffer. To estimate the quantity and quality (in terms
25 µl reaction volume containing 1.5 µl DNA (50 ng/
of protein and RNA contamination) of isolated genomic
µ l), 2.5 µl of PCR buffer (10 x) with 15 mM MgCl 2,
DNA, spectrophotometry was performed. One microliter
Bangalore Genei, India, 1 µl of Primer (10 p moles/ µ l)
of DNA sample was loaded on to the well of Nanodrop
(MWG), 0.5 µl of dNTPs (2.5 mM) Bangalore Genei,
instrument. The concentration of DNA and absorbance at
India, 0.5 µl of Taq DNA polymerase (3U/ µ l) and 19 µl
260 nm and 280 nm were measured. To check the DNA
of sterile distilled water. Amplification was performed in
quality of isolated genomic DNA, electrophoresis was
a programmed thermocycler with initial denaturation at
done using 0.8% agarose gel prepared in 1X TBE buffer.
94°C for 5min, 35 cycles of denaturation at 94°C for 1min,
primer annealing at 38°C for 1min, extension at 72°C for
RAPD amplification
1min, and final extension at 72°C for 5 min. Amplified
Amplification of RAPD fragments was performed
products were electrophoresed on 1.5% agarose in 1 x TBE
according to Williams et al. (1990) using decamer arbitrary
buffer. The gels were stained with ethidium bromide and
primers (Operon technologies Inc, USA; SIGMA-D,
documented using gel documentation system (Bio-Rad,
USA) (Table 2). Amplification were performed in a
Hercules, California).
431