Research on genetic diversity of Chinese population of the crested ibis.

本研究的目的是测定中国濒危物种朱鹭种群的遗传多样性水平，并建立可用于监测个体动物近亲交配状况的遗传标记。为此，我们收集了死亡动物的DNA样本，并检测了样本中的遗传变异。首先，我们测定了线粒体DNA(MtDNA)的部分核苷酸序列，并分析了朱鹭与其近缘物种的系统发育关系，我们的数据表明朱鹭与红唇朱鹭、光彩朱鹭的亲缘关系最近。然而，我们没有发现线粒体DNA序列在个体之间的变异。另一方面，扩增片段长度多态(AFLP)方法在个体水平上发现了变异。在AFLP实验中，我们使用了EcoRI和MseI限制性内切酶来产生DNA片段。在AFLP的最后一步，我们用EcoRI-限制性引物和MseI-限制性引物对筛选出的DNA片段进行扩增，用12组选择性引物来限制DNA条带的数目。结果表明，如果能鉴定出更多的AFLIP标记，AFLP技术将适用于朱鹭群体的系谱分析。在样本群体中也发现了微卫星和非编码序列等其他核标记，但到目前为止还没有发现样本之间的变异。我们还对部分主要组织相容性复合体基因座进行了测序，但未能发现遗传变异。这些数据表明，中国朱鹭种群处于极端近亲繁殖状态，遗传多样性水平大幅下降，但仍存在一些变异。

The objective of this study is to measure the level of genetic diversity of a population of the crested ibis, an endangered species, in China, and to establish genetic markers that can be utilized for monitoring inbreeding status of individual animals. For this purpose, we collected DNA sapmles from dead animals and examine genetic variations among the samples. First, we determined a partial nucleotide sequence of mitochondrial DNA (mtDNA), and analyzed phylogenetic relationship between the crested ibis and its related species, Our data shows that the crested ibis is most closely related to Plegadis falcinellus, Glossy Ibis. We, however, found no variation in mtDNA sequence among individuals. On the other hand, variations at individual level were found by the method of amplified fragment length polymorphism (AFLP). In AFLP experiment, we used EcoRI and MseI restriction endonucleases to generate DNA fragments. Then we used EcoRI-restricted primer and MseI-restricted primer to amplify selected DNA fragments At final step of AFLP, we used 12sets of selective primers to limit the number of DNA bands in the analysis. Two sets of the primers gave different fragment pattern for individual samples, The result suggest that AFLP would be applicable to determine pedigree structures in the crested ibis population if more AFLIP markers are identified. Other nuclear markers such as microsatellite and non-coding sequences were also identified in the sample population, however, no variation was found among samples so far. We also sequenced a part of major histocompatibility complex loci, but failed to find genetic variations. Those data suggest that the population of the Chinese crested ibis is tinder extreme inbreeding condition and the level of genetic diversity is greatly reduced, although some variations still exist.