【作者】 刘畅；

【导师】 刘宝；

【作者基本信息】 东北师范大学 ， 遗传学， 2015， 博士

【摘要】 中间偃麦草(Thinopyrum intermedium,2n=42)是禾本科偃麦草属的多年生植物,是小麦的野生近缘物种之一。中间偃麦草具有很多优良性状,可为小麦育种提供有益基因,也是偃麦草属中最早同小麦杂交成功的物种。小麦与中间偃麦草杂交后代衍生出很多新的品系,如双二倍体,异附加系和异代换系等。本研究利用荧光原位杂交(FISH)和基因组原位杂交(GISH)相结合的方法,对中间偃麦草,小偃麦双二倍体中1—中5,以及异附加系TAI系列进行了核型的鉴定和分析。首先,以簇毛麦和拟鹅观草的基因组DNA为探针,粗山羊草DNA为封阻对中间偃麦草进行基因组原位杂交,成功的将其染色体组区分为三种(St,Js和J),每个基因组含有7对(14条)染色体。再结合FISH杂交结果,建立了中间偃麦草核型的标准图谱。为后续在杂交后代中追踪染色体的变化提供了细胞学基础。其次,对5个小偃麦双二倍体的核型进行鉴定,结果发现中1—中5均在普通小麦的基础上附加了12或14条中间偃麦草的染色体,其外源染色体构成分别为6Js+4St-Js+2J,6Js+4St-Js+2J,4Js+2Js-W+6St+2St-Js,2Js+2Js-W+4St-Js+2St-W+4St和2Js+2Js-W+4St-Js+6St。附加的外源染色体均是中间偃麦草的不同染色体组的染色体构成的混合基因组。在小偃麦双二倍体的形成过程中,中间偃麦草的不用基因组间有很多易位产生,也有少数小麦与中间偃麦草的易位。对附加系TAI系列核型的鉴定发现,其核型很不稳定,会发生外源染色体的丢失,或外源染色体代换了小麦的染色体,也会由于外源附加,造成小麦内部染色体不稳定从而发生代换或易位。综合双二倍体和附加系每种类型的核型,发现附加1对或2对染色体对小麦染色体稳定性造成的影响要远远强于附加一套基因组数量染色体所造成的影响;在附加系中发现了更为复杂多样的核型变化。虽然在中字号系列材料中,外源染色体类型多样,但被保留下来并遗传到附加系的种类却减少了很多,14种附加系按照外源染色体类型来分,大致分为5类,即3J,7Js,5Js,4Js-D和5St-Js。为了研究附加的外源染色体对基因表达的影响,我们选取TAI 1-4的三种核型进行转录组测序。通过数据分析发现附加一条或是两条外源染色体对于总体基因表达水平影响不大。通过SNP鉴定外源基因,发现外源基因在附加系中表达水平很低,不足以引起基因的差异表达;而原来这部分基因处于中间偃麦草中时的表达水平明显高于附加系。对小偃麦双二倍体及异附加系混合基因组构成的深入研究,解析小偃麦系列物种类型形成的遗传基础,对于加深对植物界多倍体新物种形成的规律认识具有重要意义。进一步结合组学研究方法,对外源染色体携带基因进行更为细致的分析,为解析小偃麦提供新的方法和思路,将推动小偃麦系列的遗传和应用研究的深入开展。更多还原

【Abstract】 Thinopyrum intermedium(2n=42), which is one of the wild relative species of wheat, is a perennial plant of Elytrigia. Th. intermedium possesses numerous agriculturally desirable characteristics, and thus may provide useful genes for wheat genetic improvement. In Elytrigia, Th. intermedium is actually the first successful case to hybridizing with wheat, and many kinds of new germplasm lines were developed from the progenies of common wheat and Th. intermedium, including amphidiploids, addition lines and substitution lines etc. In the present study, fluorescence in situ hybridization(FISH) and genome in situ hybridization(GISH) were used to investigate the karyotypes of Th. intermedium, amphidiploids of wheat(Triticum aestivum) – Th. intermedium(Trititrigia) and the “TAI” series of alien addition lines.Based on GISH using genomic DNA of Pseudoroegneria strigosa and Dasypyrum villosum as probes and DNA from Aegilops tauschii as a block, Th. intermedium genome was divided into three subgenome sets(St,Js and J) and 14 chromosomes(7 pairs) per set. Combination of the results of FISH and GISH, the ideogram of Th. intermedium was established, which has laid an indispensible for further studies.Karyotypic analysis of the five Trititrigia amphidiploids showed that, Zhong1-Zhong5 all had the whole set of wheat chromosomes plus 12-14 Th. intermedium chromosomes, and the constitutions of alien chromosomes were 6Js + 4St-Js + 2J,6Js + 4St-Js + 2J,4Js + 2Js-W + 6St+ 2St-Js,2Js + 2Js-W + 4St-Js + 2S-W + 4St and 2Js + 2Js-W + 4St-Js + 6St. The alien chromosomes in each amphidiploid were all from different subgenomes of Th. intermedium. During the formation of Trititrigia amphidiploids, many translocations have taken places between different genomes of Th.intermedium, and some between common wheat and Th.intermedium.The two sets of alien addition lines(the TAI series) showed highly unstable karyotypes, including loss of alien chromosomes, substitution of wheat chromosomes by alien chromosomes, and substitution and translocation between different chromosomes of wheat. In contrast, we found that the karyotypes of amphidiploids were more stable than those of the addition lines. Changes of karyotypes in addition aliens were more complex and diverse. According to the alien chromosomes from Th.intermedium, which were structurally modified in the Trititrigia amphidiploids, the two sets of alien addition lines were divided into only five types, 3J,7Js,5Js,4Js-D and 5St-Js.For analysis the impact of the alien chromosomes on gene expression, three karyotypes of TAI I-4 were chosen for transcriptome sequencing. The data showed that there were no dramatic changes between wheat and alien addition lines. Alien genes were identified by SNP, and we found that these genes expressed at a much higher level in Th.intermedium than in TAI I-4, suggesting general suppression of gene activity from alien chromosomes by the wheat genome.Further analysis of the mechanisms leading to formation of mixed genomes and the genetic basis of the Tritirigia amphidiploids may provide novel insights into our current understanding of polyploidy speciation in plants.更多还原