关键词: 国兰, 种质资源, 遗传多样性, SRAP标记, 分子身份证

Abstract: 【Objective】 The genetic diversity of Chinese cymbidium germplasms was studied, meanwhile, the molecular fingerprinting and ID card were established in order to provide technique support for their identification at molecular level, and to lay the root for their exploitation, conservation, utilization and innovation in the future. 【Method】 139 samples of Chinese cymbidium gemplasm containing cultivated and wild species were collected from south China and its neighborhood. Their genomic DNA was extracted by the modified method of CTAB. 208 pair SRAP primers was from random combinations with 13 pieces F-primer and 16 pieces R-primer, which each pair of SRAP primers was initially screened for amplification of a specific product from genomic DNA of two cultivars, one from ‘Songmei’ and the other from ‘Daxun’. The PCR amplification products were separated on 6% nondenaturing polyacrylamidegel electrophoresis. The electrophoresis pattern was artificially analyzed, and total bands, polymorphic bands and percentage of polymorphic bands(PPB) were counted. The polymorphism information content(PIC) were counted according to Botstein’s formula. The observed number of alleles (Na), effective number of alleles (Ne), Nei’s genetic diversity diversity index (H) , and Shannon’s information index (I) all were estimated by software POPGENE32. The UPGMA clustering analysis was performed by software NTSYS-pc2.10e, and attained the genetic similarity coefficient value. The method of primer combinations was used to construct a digital fingerprinting of 139 samples of Chinese cymbidium germplasm. 【Result】 17 pair polymorphic primers were screened from 208 pair SRAP primers, and a total of 489 bands were amplified by these primers, of which 484 bands were polymorphic and PPB was 98.89%, observed number of alleles (Na), the effective number of alleles (Ne), Nei’s genetic diversity diversity index (H) , and Shannon’s information index (I) were respectively 2.00, 1.49, 0.30, 0.45. The genetic similar coefficient of 139 Chinese cymbidium ranged from 0.51 to 0.91, and the cluster analysis based on UPGMA showed that these germplasm can be divided into six major groups at similarity coefficient of 0.70. All of the samples could be effectively distinguished by different primer combinations. The specific molecular identity for 139 samples of Chinese cymbidium germplasm were established with 99.99% probability of confidence. However, Chinese cymbidium germplasm can be divided into four sections according to their genetic relationship, section one of which is C. goeringii germline, only consists of one species, section two mainly includes C. ensifolium and C. sinense germline, section three consists of C. faberi, C. kanran andhybrid species, section four includes C. longibractium and C. lianpan. In terms of genetic relationship among them, section four is the nearest with section three, section two comes second, is farthest with section one.【Conclusion】 A higher level of genetic diversity exists in Chinese cymbidium germplasm examined, it was unique and efficient that the molecular identity system of 139 samples was first established based on 17 pair primer combinations of SRAP. The SRAP technique can provide a powerful tool for Chinese cymbidium germplasm identification.

Key words: Chinese cymbidium, germplasm, genetic diversity, SRAP marker, molecular ID card