Analysis Of ISSR On Genetic Diversity Of The Varieties Of Medicago Sativa.L

By the usage of ISSR analyzing technique, the study probes into genetic diversities among 30 alfalfa varieties, which work as experimental material and mainly come from China, America and Australia. Major results were as follows:(1) The establishment of an optimized PCR reaction system for ISSR analysis: PCRwas performed in a 25ul reaction mixture with 20-100ng DNA, 0.5μmol/L primer, 0.2 mmol/L dNTP, 0.5U Taq DNA polymerase, 2.5 mmol/L Mg²⁺, 2μl 10×buffer. The reaction process went as following: the temperature profile used for PCR was 94°C for 2 minutes, followed by 35 cycles of 94°C for 30 seconds ,50-55°C for 30 seconds (the temperature depends on the Tm value of primer), 72°C for 2 minutes, and was terminated with a 10- minute DNA extension step at 72°C.(2) The primer sieving of alfalfa ISSR and amplification of PCR: in 30 experimentalvarieties, 109 DNA bands were amplified by 10 ISSR primers, in which 70 bands were polymorphic, polymorphic ratio (PPB) was 64.22% .10.9 bands were amplified by every primer.and among all bands amplified, 105, with average polymorphic ratio (PPB) as 64.22%.it can be seen from the widely distributing in worldwide area, and adapting to various climate and soil condition, alfalfa have strong adaption and genetic ability,potential of breeding amending.(3) The molecular cluster of alfalfa variety about ISSR mark: based on the ISSR markbands and the summarized information separately, the UPGMA cluster analysis was carried on, the results indicated that: the 30 varieties were divided into two groups, one group was consist of 16 alfalfa varieties coming from the northern area, other group was composed of 13 alfalfa varieties coming from overseas and the southern area. The result was also shown that there was relation of heredity of alfalfa with its dormancy index..