【作者】 黄燕；

【导师】 周永红；

【作者基本信息】 四川农业大学 ， 植物学， 2009， 硕士

【摘要】 猬草属(Hystrix Moench)是禾本科(Poaceae)小麦族(Triticeae)的一个多年生小属,全世界约10余种,主要分布于北美和中亚、西亚。Moench(1794)把Linneus(1753)描述的披碱草属(Elymus L.)物种——Elymus hystrix L.,因为其颖强烈退化甚至缺失的特点,建立了猬草属(Hystrix Moench),其模式种为Hystrix patula Moench。迄今为止,报道的Hystrix物种约11个种,中国有3种。Church(1967)报道Hy.patula与加拿大披碱草复合群(Elymus canadensis L.complex)物种具有较近的亲缘关系。Dewey(1984)报道了Hy.patula具有StH染色体组,并根据染色体组分类的原则,将Hystrix物种放在Elymus中。Jensen et al.(1997)报道了Elymus coreanus(Hy.coreana)和E.californicus(Hy.californica)具有与赖草属(Leymus Hochst.)相同的NsXm染色体组,并把这两个物种组合到Leymus中,使得Hystrix的分类更为复杂。因此,Hystrix属的界限和物种划分一直处在争议之中,其主要分歧是将这些物种置于Hystrix?还是作为Elymus的一部分?以及还是放在Leymus中?东北猬草(Hystrix komarovii(Roshev.)Ohwi)是猬草属的一个四倍体(2n=4x=28)物种。Ellenskog-Staam et al.(2007)根据GISH和Southern杂交结果,报道Hy.komarovii可能具有St和H染色体组,是StH的一种变异类型。然而,Zhang et al.(2009)根据nrDNA ITS序列分析认为Hy.komarovii可能含有与Hy.coreana和Hy.duthiei相似的NsXm染色体组,而不具有Hy.patula的StH染色体组。本研究通过基因组特异的RAPD标记、基因组原位杂交(GISH)和DNA序列分析,探讨东北猬草的染色体组组成。主要结果如下:1.对东北猬草和其他4个猬草属物种以及8个近缘属物种进行基因组特异性RAPD标记研究。选用5个基因组特异的RAPD引物(St、H、Ns、E~e和E~b)进行PCR扩增,结果显示:(1)Hy.patula具有St和H基因组特异的RAPD标记,而Hy.komarovii和其它猬草属物种没有St和H基因组特异的RAPD标记;(2)Hy.komarovii与Hy.duthieissp.duthiei、Hy.duthiei ssp.longearistata及Hy.coreana一致,具有Ns基因组特异的RAPD标记,而Hy.patula没有Ns基因组特异的RAPD标记;(3)猬草属和赖草属物种均没有E~e和E~b基因组特异的RAPD标记。该实验结果表明Hy.komarovii可能具有Ns染色体组而不具有St和H染色体组。2.对东北猬草和其他4个猬草属物种进行单色基因组原位杂交(GISH)分析。利用具不同基因组的4个二倍体物种分别作为探针DNA和封阻DNA:Pseudoroegneriaspicata(St)、Hordeum bogdanii(H)、Psathyrostachys juncea(Ns)、Lophopyrum elongatum(E~e)。结果表明:(1)Hy.patula由StH染色体组组成;(2)Hy.duthiei ssp.duthiei、Hy.duthiei ssp.longearistata及Hy.coreana具有新麦草属的Ns染色体组,另一染色体组与E~e染色体组存在重复片段同源;(3)Hy.komarovii的28条染色体中,14条显示强烈的新麦草属的Ns探针信号,14条显示或强或弱的E~e探针信号。结果表明Hy.komarovii具有一组来自新麦草属的Ns染色体组,另一染色体组与Lophopyrum的E~e同源性很高。3.对东北猬草和其他4个猬草属物种及其9个近缘属物种的细胞核DNA ITS序列进行了系统发育关系分析。在系统发育树中,Hy.komarovii、Hy.duthiei ssp.duthiei、Hy.duthiei ssp.longearistata及Hy.coreana与新麦草属和赖草属物种聚成一支;Hy.patula与其他披碱草属物种聚成一支。ITS序列分析结果表明Hy.komarovii可能具有赖草属的NsXm染色体组,支持细胞学和GISH分析结果。4.根据基因组特异的RAPD标记、GISH和nrDNA ITS序列分析结果,表明:东北猬草含有一组Ns染色体组,另一组染色体组与Lo.elongatum的E~e染色体组同源性很高;东北猬草不含有披碱草属的StH染色体组。更多还原

【Abstract】 Hystrix Moench is a small perennial genus in the tribe Triticeae(Poaceae).It was established by Moench(1794) according to the distinct morphological character of lacking glumes,or long setaceous awm-shaped ones,if present.The type species is Hystrix patula Moench.Since then,eleven species have been included in Hystrix,and three of them growing in China.Cytologically,Church(1967) reported that Hy.patula had a close affinity to species of the Elymus canadensis L.complex.Dewey(1984) reported that Hy.patula had the StH genomes and included Hystrix in the genus Elymus L.according to genome classification. Jensen et al.(1997) reported that Elymus coreanus(Hy.coreana) and E.californicus(Hy. californica) have the same genome constitution of Leymus(NsXm),and included them in the genus Leymus Hochst.Thus,the taxonomy of Hystrix becomes complicated.The definition of Hystrix and its precise taxonomic rank are still under discussion.The main dispute is whether included the species of Hystrix in Hystrix,or regard it as a part of Elymus or Leymus.Hystrix komarovii(Roshev.) Ohwi is a tetraploid species(2n=4x=28) in Hystrix. Ellenskog-Staam et al.(2007) reported that Hy.komarovii likely had a variant of the StH genome based on the results of GISH and Southern genomic hybridization.However,Zhang et al.(2009) suggested that Hy.komarovii might have similar NsXm genomes to Hy.coreana and Hy.duthiei,but not the StH genomes of H.patula based on the nrDNA ITS sequences.In order to investigate the genome constitution of Hy.komarovii,genomic-specific RAPD markers,genomic in situ hybridization(GISH) and nrDNA ITS sequences analysis of Hy. komarovii,the other four Hystrix species and the related species were carried out in this study. The main results are as follows:1.Five genomic-specific RAPD markers representing the St、H、Ns、E~e and E~b genome respectively were utilized to determine the genome relationships among Hy.komarovii and the related species.The results showed that:(1) the St and H genomic-specific RAPD markers presented in Hy.patula and other Elymus species containing St and H genome,but absent in Hy.komarovii,Hy.duthiei ssp.duthiei,Hy.duthiei ssp.longearistata and Hy. coreana;(2) the Ns genomic-specific RAPD marker presented in Hy.komarovii as well as in Hy.duthiei ssp.duthiei,Hy.duthiei ssp.longearistata and Hy.coreana,but absent in Hy. patula;(3) the E~e and E~b genomic-specific RAPD markers absent in either Leymus or Hystrix species.The results suggested that Hy.komarovii might contain the Ns genome,but absent the St and H genome.2.Four genomic DNA probes containing St,H,Ns,E~e genome,respectively,were used in genomic in situ hybridization(GISH) for detecting the presence of these four genomes in Hy.komarovii and the other Hystrix species.The results showed that:(1) Hy.patula had the St and H genome;(2) Hy.duthiei ssp.duthiei,Hy.duthiei ssp.longearistata and Hy.coreana had the Ns genome of Psathyrostachys and the other genome shared one or a few repetitive sequences with the E~e genome;(3) the 14 chromosomes showed strong green fluorescence when 28 chromosomes of Hy.komarovii were hybridized with the probed Ns-genome DNA of Psathyrostachys,while the 14 chromosomes showed strong or weak green hybridization signals when the chromosomes of Hy.komarovii were hybridized with the probed E~e-genome DNA of Lophopyrum elongatum.The results suggested that Hy.komarovii had the Ns genome of Psathyrostachys,and the other genome of Hy.komarovii had high homology with the E~e genome of Lophopyrum.3.To estimate the phylogenetic relationships of Hy.komarovii and the related species, nuclear ribosomal internal transcribed spacer(ITS) of 14 taxa in Triticeae containing St,H, Ns,E~e,E~b,StH and NsXm genomes were analyzed.In the ITS tree,Hy.komarovii,Hy. duthiei ssp.duthiei,Hy.duthiei ssp.longearistata and Hy.coreana formed one group with the species from Psathyrostachys and Leymus,which indicated a close phylogenetic relationship among these species.Hy.patula was claded with other two Elymus species.The results indicate that Hy.komarovii might had the NsXm genomes of Leymus,which is in agreement with those of cytology and GISH.4.According to the results of genomic-specific RAPD assay,GISH,and nrDNA ITS sequenes analysis in the present study,it is concluded that Hy.komarovii had a set of Ns genome and the other genome had high homology with the E~e genome of Lophopyrum.Hy. komarovii absent the StH genomes of Elymus.更多还原