送春与多花兰种间杂交后代的ISSR分析

ISSR analysis of interspecific hybrids descendants of Cymbidium cyperifolium var. szechuanicum and C. floribundum

关键词:  送春, 多花兰, 种间杂种, ISSR分子量标记, 孟德尔分离, 聚类分析

Key words:  Cymbidium cyperifolium var. szechuanicum, Cymbidium floribundum, interspecific hybrid, ISSR markers, Mendelian segregation, cluster analysis

摘要:

该文使用简单重复序列间(ISSR)分子标记,对送春与多花兰种间杂交后代进行了研究。结果表明:从80个ISSR引物中筛选出14个扩增效果稳定的ISSR引物,对两亲本和59个F1代个体进行了ISSR扩增,得到107个扩增位点,扩增的片段大小位于90～2 100 bp之间,平均每个引物扩增7.64条条带,得到11种类型的带。ISSR标记在送春 × 多花兰的F1代中表现出一定的多态性,分离频率为44.86%,分离位点有83.33%符合孟德尔11或31的分离规律,产生偏孟德尔分离的位点占12.50%,余下的4.17%属于特殊分离带型。可能导致后代变异的位点为偏孟德尔分离的6条带、缺失的8条带或新生成的2条带。聚类图中父本和母本与F1代个体间的遗传距离较远,59个杂交后代先聚集成一组,再同母本相聚为一组,最后才同父本聚在一起,59个杂种均偏母本型。送春与多花兰的杂交后代在植株形态、染色体、遗传物质方面都具备双亲特点,61个个体间的ISSR分子量标记结果和植株形态学特征都说明,59个F1代杂种包含送春和多花兰的遗传特性是真杂种; F1代杂种既有双亲的互补特征带,又有双亲的重组片断即产生新的特异带,这说明送春与多花兰的杂交后代具有遗传变异的特点。该研究结果可以有效地对杂交后代进行定向选择,为兰花的杂交育种提供了分子依据。

Abstract:

Interspecific hybrids descendants of Cymbidium cyperifolium var. szechuanicum and C. floribundum were studied by using ISSR marker. Fourteen ISSR primers were selected from 80 ISSR primers, which were used to amplify 61 individuals including paternal, maternal and 59 F1 progeny, 107 DNA fragments were produced by using these ISSR primers to amplify, every primer had 7.64 bands in average. The length of amplified DNA fragments ranged from 90 bp to 2 100 bp, and there were 11 kinds of bands. The results showed that the polymorphism of ISSR marker in the F1 progenies was high, and the segregation locus was 44.86%. And 83.33% segregation loci accorded with the segregation patterns of Mendelian segregation model with the segregation ratio nearly 1:1 or 3:1. 12.50% segregation loci deviated from Mendelian segregation model, 4.17% segregation loci belonged to special bands. Six deviated from Mendelian segregation model bands, eight deficiency bands and two new genetic character bands would cause some new variation. Cluster analysis showed that the genetic distances of both parental and 59 F1 progeny is further, fifty-nine F1 progeny in one group, then fifty-nine F1 progeny and female plant were in one group, at last 59 F1 progeny and female plant and male plant were in one group, this result indicated that 59 hybrids were partial female type. The hybrids of Cymbidium cyperifolium var. szechuanicum and C. floribundum had some common characteristics of their parents, such as leaves, chromosomes and hereditary material. Sixty-one materials were identified by ISSR markers and morphological characters that 59 F1 progeny obtained the genetic characteristics of Cymbidium cyperifolim var. szechuanicum and C. floribundum, so they were true hybrids. Fifty-nine F1 hybrids had mutually complementary bands of both parents, the F1 hybrids also produced some new specific bands which were recombinant fragment of parents. This experiment showed that 59 F1 progeny had both genetic and variation. Therefore, ISSR markers can be used as an effective molecular technique for directional selection of the interspecific hybrid and the study of orchid cross-breeding.