RAPD And ISSR Analysis Of Genetic Diversity And Genetic Relationships Of Tobacco

In order to provide new evidence for the study of origin and evolution of (?)obacco, genetic diversity and genetic relationships among 30 accessions of Nicoliana, including 3 wild species, 3 burley tobaccos, 5suncured tobaccos, 2 aromatic tobaccos and 17 flue-cured tobaccos, collected from China, America, Canada, Peru and Australia, were investigated with the technique of RAPD andISSR analysis. Major results are as follows:(l)An optimized PCR reaction systerm for ISSR analysis was established:PCR was performed in a 25μl reaction mixture with 20-60ng DNA, 150μM/L of each dNTP, 0.18μM/L ISSR primer, 1.5U Taq polymerase, 1 × Buffer [10mmol/Ltris-HCL ( PH8.0 ) , 50mmol/L KCL , 1.5mmoI/LMgCL₂, 0.08%NonidetP40].The temperature profile used for PCR was 94℃ for 5min,followed by 41 cycles of 94°C for 45s, 54°C for lmin30s, 72°C for lmin30s,and was terminated with a 7min DNA extension step at 72°C.(2)336 and 309 DNA bands were amplified with 16 RAPD and 16 ISSR primers, among which,315(93.75%)and 288(93.20%)were polymorphic,with 21 and 19.31 bands per primer in average.(3) UPGMA cluster analysis was carried out using software Bio ID++.Two dendrograms were constructed based on the RAPD , ISSR data respectively and their integrated data. According to the trees the accessions could be clustered into three groups(one of 27 cultivars from different areas,one of two wild species from Australia and a single wild specy from Peru)at the levels of...