M.S. Alam
Department of Genetics and Plant Breeding, Bangladesh Agricultural University, Mymensingh, Bangladesh.
S.N. Begum
Senior Scientific Officer
Plant Breeding Division, Bangladesh Institute of Nuclear Agriculture, Mymensingh, Bangladesh.
R. Gupta
Scientific Officer
Bangladesh Institute of Nuclear Agriculture, Sub Station, Khagrachari, Bangladesh.
S.N. Islam
Department of Genetics and Plant Breeding, Bangladesh Agricultural University, Mymensingh, Bangladesh.
Crop Diversity, Characterization, Genetic distance, Genetic Iidentity, Polymorphic Loci
Biotechnology Laboratory, Biotechnology Division, Bangladesh Institute of Nuclear Agriculture, Mymensingh
Variety and Species
Thirty five rice genotypes including 33 landraces (i.e. Hati bajore, Malagoti, Kuchra, Enghi, Jamai naru, Hari, Dakh shail, Moina moti, Marish shail, Patnai, Bhute shallot, Kute patnai, Moghai balam, Sada gotal, Khak shail, Jota balam, Khainol, Hamai, Sylhet balam, Mota aman, Ghigoj, Piarjat, Lal biroi, Lalanamia, Golapi, Asam binni, Kakua binni, Ledra binni, Rotisail, Genggeng binni, Jolkumri, Mowbinni and Bogi) and 1 HYV of Bangladesh (i.e. Binadhan-8) and 1 Indian landrace (i.e. Nona bokhra) were used for genetic diversity analysis through RAPD markers. In order to carry out RAPD analysis, young, vigorously growing fresh leaf samples were collected from 25 days old seedling of each genotype and used as the source of genomic DNA. DNA was extracted from the leaves of each genotype using the Cetyl Trimethyl Ammonium Bromide (CTAB) mini-prep method. The simplified mini scale procedure for DNA isolation in PCR analysis developed at IRRI was followed. Confirmation of the DNA was done through electrophoresis on a 0.8% agarose gel and DNA quantification was through the spectrophotometer’s (spectronic@ GenesisrTM). Ten primers of random sequence were screened for amplification of the DNA sequences. Primers resulting in faint or irreproducible bands were excluded from subsequent analysis. A final subset of seven primers exhibiting good quality banding patterns and sufficient variability from where finally three primers were selected for further analysis. PCR reactions were performed on each DNA sample in a 10μl reaction mix containing 1 μl Ampli Taq polymerase buffer (10X), 2.5 μl Primer (10 μM), 1 μl dNTPs (250 μM), 0.2 μl Ampli Taq DNA polymerase and 3.3 μl sterile deionized water. Two μl genomic DNA was added and finally, total volume was made 10 μl. DNA amplification was performed in an oil-free thermal cycler. The reaction mix was preheated at 940C for 3 minutes followed by 40 cycles of 1 min denaturation at 940C, 1 min annealing at 540C and elongation or extension at 720C for 2 minutes. After the last cycle, a final step for 7 minutes at 720C to allow complete extension of all amplified fragments. After completion of cycling program, reactions were held at 40C. Electrophoresis was carried out in 0.5 X TBE buffer on a 1.5 % agarose gel and amplified fragments were visualized by staining with ethidium bromide.
ISSN: 2224-0616
Int. J. Agril. Res. Innov. & Tech. 4 (1): 77-87, June, 2014 Available online at http://www.ijarit.webs.com
Journal