Genetic effects of Agropyron cristatum 2P chromosome translocation fragments in a wheat background

Agropyron cristatum (2n=4x=28, PPPP) is a wild relative of common wheat which contains a large number of desirable genes that can be exploited for wheat improvement. Wheat–A. cristatum 2P alien translocation lines exhibit many desirable traits, such as small flag leaves, a high spikelet number and density, and a compact plant type. An agronomic trait evaluation and a genetic analysis were carried out on translocation lines and backcross populations of these lines carrying different translocation fragments. The results showed that a translocation fragment from 2PT-3 (2PL) reduced the length of the flag leaves, while translocation fragments from 2PT-3 (2PL) and 2PT-5 (2PL (0.60–1.00)) reduced the width of the flag leaves. A translocation fragment from 2PT-13 (2PS (0.18–0.36)) increased the length and area of the flag leaves. Translocation fragments from 2PT-3 (2PL) and 2PT-8 (2PL (0.86–1.00)) increased the density of spikelets. Translocation fragments from 2PT-7 (2PL (0.00–0.09)), 2PT-8 (2PL (0.86–1.00)), 2PT-10 (2PS), and 2PT-13 (2PS (0.18–0.36)) reduced plant height. This study provides a scientific basis for the effective utilization of wheat–A. cristatum translocation lines.     

基于小麦SNP芯片对簇毛麦6V#2和6V#4染色体及其与小麦6A、6D染色体的多态性分析

【目的】利用大数据比较2条不同的簇毛麦6V（#2和#4）染色体及其与小麦6A、6D染色体间DNA水平上的差异,为小麦-簇毛麦靶向易位的精准设计育种提供依据。【方法】以6V#4（6D）异代换系RW15为父本和6V#2（6A）异代换系南87-88为母本进行杂交,获得F₂分离群体,利用6V#4S/6V#2S/6AS/6DS/6VL特异分子标记检测F₂植株,筛选新类型的代换系,并用分子标记结合基因组原位杂交（genomic in situ hybridization,GISH）对新类型代换系进行确认,再利用小麦55K芯片中的6A、6D探针,对新代换系及其双亲南87-88和RW15进行分析;结合660K芯片6A、6D探针对2份簇毛麦的SNP分析结果,筛选6V特异SNP。【结果】GISH分析表明,19EL124和19EL134的体细胞染色体数2n=42,分别携带2条完整的外源染色体;分子标记鉴定结果表明,19EL124含有6V#4S/6DS特异标记带,缺失了6V#2S/6AS特异带,而19EL134含有6V#2S/6AS特异标记带,缺失了6V#4S/6DS特异带;19EL124和19EL134都含有6VL的特异带,证明19EL124为6V#4（6A）异代换系,19EL134为6V#2（6D）异代换系。55K芯片检测结果表明,异代换系中关键染色体探针的检测效率显著低于其他染色体,且对同类型异代换不同系的检测效率也有所不同。1 177个6A探针中,63.21%不能对6A代换系南87-88分型,68.90%不能对6A异代换系19EL124分型,22.51%检测到6V#2和6V#4间的多态性,其中88个只能检测到6V#4染色体,而155个只能检测到6V#2染色体;479个6D探针中,49.48%不能对6D异代换系RW15分型,53.44%不能对6D代换系19EL134分型,16.70%检测到6V#2和6V#4间的多态性,其中23个只能检测到6V#2染色体,42个只能检测到6V#4染色体。整合55K和660K芯片的共有探针,分别从395个6A、231个6D探针筛选获得簇毛麦6V特异的SNP标记22个和15个,其中3个可在6V#2和6V#4染色体间显示多态性。【结论】小麦染色体的缺失与外源染色体的替换,使相应染色体探针的检测效率大幅降低,NA分型比例极大增加,且多数NA分型在2条不同的外源染色体间显示多态;相同探针对2条外源染色体的检测效率不同,小麦6A探针可以更好地检测6V#2,而小麦6D探针可更好检测6V#4;在簇毛麦与异代换系6V染色体的一致性分型中,筛选获得簇毛麦6V特异的SNP标记37个。     

一个耐盐小麦-多枝赖草二体异附加系外源染色体的鉴定

 【目的】对附加系Line15进行耐盐性验证，并了解其外源多枝赖草染色体的遗传背景。【方法】利用含0.4%（w/w）NaCl的人工模拟盐池进行耐盐性鉴定，并对Line15根尖细胞、花粉母细胞的染色体数目进行检测，进而利用荧光原位杂交和微卫星标记技术对其遗传物质进行进一步鉴定。【结果】小麦-多枝赖草杂交后代Line15为一个具有较高耐盐特性的材料，其染色体组成为2n=44=22 II，属于一个小麦-多枝赖草二体异附加系。根据SSR检测结果表明，Line15附加的一对多枝赖草染色体与小麦的第2部分同源群长臂、第3部分同源群着丝粒和第7部分同源群短臂密切相关。【结论】对小麦-多枝赖草二体异附加系Line15进行了生理、细胞和分子水平的系统检测，较全面地揭示了Line15耐盐性的具体来源。     

Development and characterization of a novel common wheat–Mexico Rye T1DL·1RS translocation line with stripe rust and powdery mildew resistance

Rye (Secale cereale L., 2n=2x=14, RR) is a significant genetic resource for improving common wheat because of its resistance to multiple diseases and abiotic-stress tolerant traits. The 1RS chromosome from the German cultivated rye variety Petkus is critical in wheat breeding. However, its weakened disease resistance highlights the need to identify new resources. In the present study, a novel derived line called D27 was developed from common wheat and Mexico Rye. Cytological observations characterized the karyotype of D27 as 2n=42=21 II. Genomic in situ hybridization indicated that a pair of whole-arm translocated Mexico Rye chromosomes were inherited typically in the mitotic and meiosis stages of D27. Experiments using fluorescence in situ hybridization (FISH) and gliadin electrophoresis showed that D27 lacked wheat 1DS chromosomes. They were replaced by 1RS chromosomes of Mexico Rye, supported by wheat simple-sequence repeat markers, rye sequence characterized amplified region markers, and wheat 40K SNP array analysis. The wheat 1DS chromosomes could not be detected by molecular markers and wheat SNP array, but the presence of rye 1RS chromosomes was confirmed. Agronomic trait assessments indicated that D27 had a higher tiller number and enhanced stripe rust and powdery mildew resistance. In addition, dough properties analysis showed that replacing 1DS led to higher viscosity and lower dough elasticity in D27, which was beneficial for cake making. In conclusion, the novel cytogenetically stable common wheat–Mexico Rye T1DL·1RS translocation line D27 offers excellent potential as outstanding germplasm in wheat breeding programs focusing on disease resistance and yield improvement. Additionally, it can be valuable for researching the rye 1RS chromosome’s genetic diversity.     

VE161异代换系雄性不育原因的探讨及其保持

VE1 61小麦包括具有一对长穗偃麦草染色体的雄性不育异代换系、可育附加系和杂育系 ,杂育系由其代换系×附加系产生。对 VE1 61雄性不育异代换系的两种保持途径进行的比较研究表明 ,采用杂育系自交的方法较好 ,并探讨 E染色体对育性的影响     

Structural Changes of 2V Chromosome of Haynaldia villosa Induced by Gametocidal Chromosome 3C of Aegilops triuncialis

Haynaldia villosa （2n=2X= 14, VV）, a relative of wheat, plays important roles in wheat improvement mainly owing to its disease resistance. Powdery mildew resistance gene Pm21 has been successfully transferred into wheat by Cytogenetic Institute, Nanjing Agricultural University, China, and is widely used in the current wheat breeding programs. In this research, our objective is to further transfer and utilize the beneficial genes such as eye-spot resistance, yellow rust resistance, and gene of the tufted bristles on the glume ridge （a remarkable morphology） mapped on 2V of Haynaldia villosa. A disomic addition line with gametocidal chromosome 3C ofAegilops triuncialis added in Norin-26 was crossed to the wheat-H, villosa disomic substitution 2V（2D） and the hybrid F1 was then self-crossed. Chromosome C-banding, genomic in situ hybridization （GISH）, and meiotic analysis in combination with molecular markers were applied to detect the chromosome variations derived from hybrids Fz and F3. To date, four translocations including one small segmental translocation T6BS·6BL-2VS, two whole arm translocations （preliminarily designed as T3DS·2VL and T2VS.7DL） and one intercalary translocation T2VS·2VL-W-2VL, one deletion Del. 2VS·2VL-, one monotelosomic Mt2VS, and one isochromosome 2VS·2VS line have been developed and characterized. One wheat SSR marker Xwmc25.120 tagging 2VS and one wheat STS marker NAU/STSBCD135-1 （2BL） tagging 2VL were successfully used to confirm the alien chromosome segments involved in the seven lines. The tufted bristles on the glume ridge appeared in lines T2VS-7DL, Mt2VS, 2VS-2VS as well as the parent DS2V（2D）, whereas in T3DS·2VL, this trait did not appear. The gene controlling the tufted bristles was located on 2VS. Gametocidal chromosome 3C ofAegilops triuncialis could successfully induce chromosome 2V structural changes.     

小偃麦种质系SN0606的形态学和细胞学分析

对从中间偃麦草与小麦品种烟农15杂种后代中选育的小麦种质系SN0606进行主要农艺性状调查和细胞学鉴定,结果表明:SN0606平均株高87.0 cm,平均穗长10.5 cm,小穗数23个,穗粒数56;其根尖细胞染色体数目为2n=42,花粉母细胞减数分裂中期Ⅰ(PMCMⅠ)染色体构型为2n=21Ⅱ;与普通小麦的杂种F1PMC MⅠ染色体构型为2n=0.78Ⅰ+20.61Ⅱ,SN0606可能是一个小麦—中间偃麦草的二体异代换系。     

利用Sod同工酶和RAPD标记鉴定小麦-中间偃麦草双体异附加系

利用Sod同工酶标记和RAPD标记对DAL1、2、3、4、5、6、7、8、9、10和11等11个小麦-中间偃麦草双体异附加系进行了鉴定。通过Sod同工酶分析，将Sod-Agil基因定位于中间偃麦草第2部分同源群染色体上，同工酶标记Sod-1Rf=0．35为异附加系DAL1、3、5、6、8、9、10和11的特有生化际记。RAPD分析结果表明，引物OPF16是特异引物，为异附加系DAL4、5、9、10和11所共有。OPF16 560是DAL5、9、10、11的特异RAPD标记，OPF16 1200是DAL4的特异RAPD标记。综合鉴定结果表明：DAL1、3、5、6、8、9、1O和11等8个异附加系中附加的是中间偃麦草第2部分同源群的染色体。DAL5、9、10、11是同一种异附加系，附加的异源染色体属于八倍体中5所携带的中间偃麦草染色体组；DAL6和DAL8为同一种异附加系：DAL2和DAL4是不同的异附加系，DAL4中附加的是不同于中2、中5所携带的中间偃麦染色体组的另外一个染色体组的染色体。应用上述遗传标记．可以快速地鉴定异附加系DAL1、3、4、5、6、8、9、10、11。     

普通小麦-簇毛麦易位系T6BS·6BL-2VS的选育

[目的]为进一步利用簇毛麦2V染色体上的有益基因,为小麦育种提供新种质。[方法]通过普通小麦-簇毛麦2V(2D)二体代换系(DS2V)与普通小麦农林26-离果山羊草3C染色体二体异附加系(DA3C)杂交,综合运用染色体C-分带、基因组原位杂交和分子标记分析,并结合性状调查。[结果]从杂种后代中选育出小麦-簇毛麦纯合易位系T6BS.6BL-2VS,性状调查发现该易位系植株护颖颖脊上有刚毛。[结论]该易位系为杀配子染色体诱发的小片段易位;簇毛麦护颖颖脊刚毛基因定位于2VS的中部至端部。     

利用PCR技术初步鉴定小麦-加州野大麦异染色体系

为快速鉴定普通小麦与普通小麦—加州野大麦双二倍体杂交、回交后代植株的染色体组成，研究小麦背景中添加的外源染色体与小麦染色体之间的部分同源关系，选用已被定位在小麦7个部分同源群21条染色体上的38个SSR引物对杂种回交后代植株进行PCR扩增。结果表明，其中27个小麦SSR引物在普通小麦与小麦—加州野大麦双二倍体间有多态性扩增，涉及4个部分同源群的11对引物，可在不同杂种回交植株中扩增出与双二倍体相同的多态带纹；根据PCR扩增和细胞遗传学分析的结果，在18个回交后代中初步鉴定出7个可能的异附加系，其中2个二体异附加系、1个端二体异附加系、2个单体异附加系和2个双单体异附加系。所选育的异附加系分别涉及第1、2、4和7部分同源群。     

Development of EST-PCR Markers for the Chromosome 4V of Haynaldia villosa and Their Application in Identification of 4V Chromosome Structural Aberrants

EST-PCR based molecular markers specific for alien chromosomes are not only useful for the detection of the introgressed alien chromatin in the wheat background, but also provide evidence of the syntenic relationship between homoeologous chromosomes. In the present study, in order to develop high density and evenly distributed molecular markers on chromosome 4V of Haynaldia villosa, a total of 607 primer pairs were designed according to the EST sequences, which were previously located in 23 different bins of wheat chromosomes 4A, 4B and 4D. By using the Triticum durum-H, villosa amphiploid and T. aestivum-H, villosa alien chromosome lines involving chromosome 4V, it was found that 9.23% of the tested primers could amplify specific bands for chromosome 4V. Thirty and twenty-six specific markers could be assigned to chromosome arms 4VS and 4VL, respectively. These 4V specific markers provided efficient tools for the characterization of structural variation involving the chromosome 4V as well as for the selection of useful genes located on chromosome 4V in breeding programs.     

人工合成的十个优良小麦种质系的细胞学和性状特点的研究

本试验对从八倍体小偃麦与普遍小麦的杂种后代中选出的10个优良小麦种质系,进行了细胞学和田间试验鉴定。结果表明,在这10个小麦种质系中,其中有2个品系为单体异附加系,1个品系为双单体异附加系,2个品系为双体异附加系。有5个品系染色体数目与普通小麦相同,但具有偃麦草的某些特点,细胞学上具有异常行为,它们可能为新的次生六倍体小麦。这些小麦种质系中,有的综合性状较好,具有较好的丰产性能,有希望在生产上直接加以利用;有的虽然综合表现欠佳,但具有突出特点,可作为种质材料在育种上利用。     

普通小麦-簇毛麦易位系T6BS·6BL-2VS的选育(英文)

[Objective] The aim of experiment was to provide a new germplasm for wheat breeding by further using desirable genes in 2V chromosome of Haynaldia villosa.[Method] Through hybridization between common wheat(Triticum aestivum)-Haynaldia villosa disomic substitution line and common wheat Nonglin26-3C chromosome of Aegilops triuncialis disomic addition line,the analysis methods such as chromosome C-banding,genomic in situ hybridization and molecular marker technique were comprehensively applied and combined characters investigation.[Result] The wheat-Haynaldia villosa translocation line(T6BS·6BL-2VS)was selected from hybrid progenies to conduct characters investigation,which found some bristles on glume ridge of T6BS·6BL-2VS.[Conclusion] The translocation line induced by gametocidal chromosome was a small segment translocation line and the gene of bristle on glume ridge of Haynaldia villosa was located between the middle and the terminal of 2VS.     

大白菜-结球甘蓝5号二体异附加系的选育及鉴定

【目的】获得稳定遗传的大白菜-结球甘蓝二体异附加系。【方法】以大白菜-结球甘蓝5号单体异附加系为材料,对其自交后代进行细胞学及SSR鉴定。【结果】37株自交后代中2n=22植株的比率为10.81%,通过对4个2n=22的植株进行核型分析及SSR鉴定,确定其为大白菜-结球甘蓝5号二体异附加系。减数分裂观察发现,大白菜-结球甘蓝5号二体异附加系后期Ⅰ虽然出现染色体落后现象,但在后期Ⅱ中染色体的分裂方式以11-11-11-11为主,其比率为69.55%。【结论】获得了大白菜-结球甘蓝5号二体异附加系,其外源染色体与甘蓝07连锁群相对应;该二体异附加系减数分裂形成的四分体以附加1条的染色体为主(n=11),有较高的遗传稳定性。二体异附加系的获得为研究芸薹属A、C基因组的亲缘关系,以及向大白菜中导入结球甘蓝的优良基因具有重要意义。     

普通小麦-华山新麦草异附加系的分子细胞遗传学研究

利用荧光原位杂交和染色体C-分带技术,对普通小麦-华山新麦草的异附加系进行了研究。荧光原位杂交结果显示:异附加系H9015-17-1-9,H9017-14-16-5都附加有2条华山新麦草的染色体。对这2个材料和华山新麦草进行染色体C-分带带型比较,初步推断,H9015-17-1-9附加的是Nh5染色体,H9017-14-16-5附加的是Nh6染色体。     

普通小麦-簇毛麦易位系T6BS·6BL-2VS的选育(英文)

[Objective] The aim of experiment was to provide a new germplasm for wheat breeding by further using desirable genes in 2V chromosome of Haynaldia villosa.[Method] Through hybridization between common wheat(Triticum aestivum)-Haynaldia villosa disomic substitution line and common wheat Nonglin26-3C chromosome of Aegilops triuncialis disomic addition line,the analysis methods such as chromosome C-banding,genomic in situ hybridization and molecular marker technique were comprehensively applied and combined characters investigation.[Result] The wheat-Haynaldia villosa translocation line(T6BS·6BL-2VS)was selected from hybrid progenies to conduct characters investigation,which found some bristles on glume ridge of T6BS·6BL-2VS.[Conclusion] The translocation line induced by gametocidal chromosome was a small segment translocation line and the gene of bristle on glume ridge of Haynaldia villosa was located between the middle and the terminal of 2VS.     

普通小麦大赖草易位系T3AS·3AL-7Lr#1S的分子细胞遗传学鉴定

大赖草对赤霉病具有较好的抗性,将大赖草赤霉病抗性基因转入普通小麦,对拓宽小麦赤霉病抗性基础有重要意义。本研究在获得抗赤霉病普通小麦-大赖草异附加系基础上,采用1 200 R ⁶⁰Co-γ射线处理小麦-大赖草二体附加系DA7Lr花粉,授予已去雄的普通小麦中国春,对其后代(M₁)种子根尖细胞有丝分裂中期染色体进行GISH分析,获得了1株具有1条普通小麦-大赖草易位染色体的植株,让其自交,对自交后代中具有2条易位染色体植株的花粉母细胞减数分裂中期Ⅰ进行观察,发现2条易位染色体形成了稳定的环状二价体,表明该植株为纯合体。利用顺序GISH-双色FISH分析,结合C-分带、小麦D基因组专化探针Oligo-pAs1-2和B基因组专化探针Oligo-pSc119.2-2,进一步鉴定出该普通小麦-大赖草易位系为T3AS·3AL-7Lr#1S,且筛选出了可追踪该易位系的3个EST-STS分子标记BE591127、BQ168298和BE591737。该易位系的育成为小麦赤霉病遗传改良提供了新种质。     

矮秆小偃麦异附加系31504的遗传分析：Ⅰ鉴定与细胞遗传

本研究利用细胞学和同工酶电泳等方法对小偃麦二体异附加系３１５０４进行了鉴定；并在不同遗传背景下．分析了异附加染色体的传递特点。结果表明，３１５０４为３Ｆ２／７Ｆ２易位的二体异附加系，其单体异染色体通过胚囊和花粉传递的频率平均分别为２８．９％和２０．２％，且与遗传背景无关。     

基于簇毛麦No.1026转录组的SSR序列分析及其PCR标记开发

【目的】 探究从前苏联引进的簇毛麦No.1026（Dv#4）的EST-SSR序列特征及其在染色体的分布,分析它们在不同簇毛麦间及与小麦间的多态性,为其进一步的研究与利用提供依据。【方法】通过Illumina HiSeq测序获得No.1026植株的转录组序列,利用MISA软件分析转录组SSR序列及特征,采用Primer 3设计SSR引物,随机合成238对引物,对小麦中国春与簇毛麦Dv#4和引自英国剑桥的簇毛麦Dv#2的基因组DNA进行扩增,在琼脂糖凝胶上分离评价扩增产物的多态性,并利用一套小麦-簇毛麦异染色体系进行扩增分析。【结果】 检测了No.1026总长62.76 Mb的转录组序列,发现10 497个SSR位点,它们分布于8 735条Unigene上。在1—6个核苷酸的重复单元中,单、双、三碱基的重复占95.85%,其中三核苷酸串联重复数量最多,占SSR总数的50.33%,而CCG/CGG基序的重复占三核苷酸串联重复的41.66%;单核苷酸重复是第二大类型,出现频率为27.13%,其中A/T重复占单核苷酸重复的74.58%。二核苷酸重复类型的数量位列第三,占SSR总数的18.39%。在238对EST-SSR引物中,88对在中国春与簇毛麦（包括Dv#2和Dv#4）之间的扩增产物显示多态性;8对只在簇毛麦和单一异染色体系扩增;4对可在簇毛麦和多个异染色体系中特异扩增;但多数可在簇毛麦中清晰扩增的引物不能在任何异染色体系中扩增,推测可能与簇毛麦基因组及染色体导入小麦过程中的变异有关;47对（19.74%）在2份不同来源的簇毛麦Dv#2和Dv#4之间的扩增产物呈现多态性。利用1对EST-SSR引物和1个EST-PCR标记检测48个簇毛麦植株,证明多态性的SSR引物可有效用于检测簇毛麦的异质性。【结果】 簇毛麦No.1026（Dv#4）的转录组中存在丰富的SSR序列,其中CCG/CGG、A/T和AG/CT等三核苷酸、单核苷酸和二核苷酸是其最主要的串联重复基序。部分EST-SSR的侧翼保守序列与单一或若干外源染色体特异相关联,据此开发特异的分子标记,可用于跟踪检测小麦背景中特异的簇毛麦染色体或染色体片段。Dv#4与Dv#2间的部分EST-SSR具有多态性,提示2份不同来源簇毛麦的表达序列间存在一定程度的遗传差异,因此,簇毛麦Dv#4值得进一步的发掘研究。