利用SSR荧光标记对野慈姑异交率的估测

对自交亲和植物交配系统进行估测,有利于了解植物的繁殖状态、自异交进化的轨迹和特定种群的自然历史或选择压力。自交亲和的野慈姑有性繁殖和克隆繁殖并存,且其花序内和无性系分株间存在雌雄花同时开放的现象即给自交带来了机会。本研究利用SSR标记估测野慈姑自然种群的异交水平,并比较分析异交率估测中荧光定量法的准确性和优势,探讨野慈姑不同自然种群微卫星位点的多态性。结果表明,从28对SSR引物中筛选出3对多态性较高的引物,其多种群水平的位点数分别是5、6、6;对1个自然种群的6棵植株共计31个果实的异交率进行估测,其整体异交率为92.87%±2.5%,揭示了野慈姑自然种群的交配系统以异交为主,且无性繁殖对后代的贡献有限;单引物水平下,荧光定量技术检测出的多态性位点数和杂合率均高于NativePAGE成像电泳,且其对异交率估测的结果也更准确;另外,从检测效率实验耗时和实验成本等方面综合分析,本研究推荐使用荧光定量技术。

An Estimation of the Outcrossing Rate in Sagittaria trifolia Using SSR Fluorescence Markers

The estimation of mating systems in self-compatible plants can help us understand the reproductive status of plants, the evolutionary trajectories between outcrossing and selfing, and the natural history or selective pressure on specific populations. Sagittaria trifolia is a self-compatible species, with both sexual and clonal reproduction. Male and female flowers sometimes co-bloom among and within inflorescences, therefore creating a chance of geitonogamy. In this study, we used SSR markers to estimate the outcrossing rate of S. trifolia in natural populations. Our experiment emphasized the polymorphism of microsatellite loci in different populations and highlighted the accuracy and advantage of the methodology of fluorescence quantification in visualizing PCR products. Results showed that 3 out of 28 primer pairs had high polymorphism, with the number of alleles at 5, 6, and 6 combining multiple populations. With the use of the three selected primer pairs, we estimated the outcrossing rate for 31 fruits from six plants in a natural population of S. trifolia, which was 92.87%±2.5%. Our study indicated that the mating system in S. trifolia was predominately outcrossing in natural populations and the contribution of clonal reproduction to offspring was very limited. We also compared two alternative methodologies in analyzing PCR products, namely Native-PAGE imaging and fluorescence quantification. The latter yielded a higher estimation on allele polymorphism, heterozygosity and therefore a more accurate outcrossing rate. After considering the cost, manpower and time collectively, we recommend employing fluorescence quantification.