Genetic Diversity Of Siraitia Grosvenorii By ISSR And RAPD Markers

Siraitia grosvenorii ,a liana species of the Cucurbitaceae endemic to Southern China, is an important economic plant There were abundant wild and cultivated Siraitia grosvenorii genotypes extensively distributed in Southern China. But,the living surrounding of wide Siraitia grosvenorii has been serious narrowed,need to conserve by scientific and effective methods. In the study,total 170 accession samples of nine populations randomly collected from Guangxi and Guangdong provinces;The genetic diversity of 130 individuals from 7 of the populations was estimated using inter-simple sequence repeat ( ISSR ) markers.,and the genetic diversity of 170 individuals from all of populations was investigated with the technique of Random Amplified Polymorphic DNA ( RAPD ) analysis. An optimized PCR reaction system for ISSR and RAPD was established: PCR was performed in a 25μL reaction mixture with 20 25 ng DNA, 200μmol/L of each dNTP, 0.5μmol/L primer,1×PCR buffer,20mmol/L MgCl₂ ,and 1 U Taq polymerase. The temperature profile used for ISSR-PCR was 94℃for 3 min,followed by 40 cycles of 94℃for 1min,52℃for 50sec,and 72℃for 2 min,and was terminated with a 7 min DNA extension step at 72℃.The temperature profile used for RAPD-PCR was 94℃for 5 min,followed by 45 cycles of 94℃for 1min,36℃for 1 min ,and 72℃for 1 min 30 sec,and was terminated with a 10 min DNA extension step at 72℃. The 15 primers employed produced a total of 111 scorable amplified fragments,of which 91 (82.0 %) are polymorphic. Genetic diversity analysis showed that Nei's gene diversity ( H_e ) was 0.248 and Shannon's genetic diversity index (I) was 0.354. Genetic differentiation mainly occurred among populations ( G_st = 0.569 ), variation among populaions is 57.40%, variation within populations is 42.60%. The correlation between genetic distance and geographic distance among the populations was not significant ( r = 0.369 ,P = 0.115 ). The 19 RAPD primers employed produced a total of 118 scorable amplified fragments,of which 98 ( 83.05% ) are polymorphic. Genetic diversity analysis showed that Nei's gene diversity ( H_e ) was 0.272 and Shannon's genetic diversity index ( I ) was 0.409. Genetic differentiation mainly occurred among populations (Gst =0.569), variation among populaions is 57.49%, varitation within populations is 42.51%. The correlation between genetic distance and geographic distance among the populations was not significant ( r = 0.370,P =0.088) . UPGMA cluster analysis was carried out using software NTSYSpc 2.11s. Two dendrograms were constructed based on the ISSR and RAPD data respectively, UPGMA analysis showed that the 130 individuals were clustered into 7 clusters corresponding to the 7 populations with the ISSR data;it showed the 170 individuals were clustered into 9 clusters corresponding to the 9 populations with the RAPD data. There was a little high correlation between the two sets of similarity parameters calculated with ISSR and RAPD data respectively ( r = 0.736,P=0.0003 ),indeicating that the two methods were consistant and able for detecting genetic diversity in grosvenorii. In summary, genetic differentiation mainly occurred among populations; it's in Jinxiu and Yongfu that there are the major regions for studing and using the grosvenorii genetic resources. Utilizing these germplasm resources as a important means of cultivating excellent foundation were provided, and the present task to our country were indicated.