A BAC-based physical map of the major autosomes of Drosophila melanogaster

We constructed a bacterial artificial chromosome (BAC)-based physical map of chromosomes 2 and 3 of Drosophila melanogaster, which constitute 81% of the genome. Sequence tagged site (STS) content, restriction fingerprinting, and polytene chromosome in situ hybridization approaches were integrated to produce a map spanning the euchromatin. Three of five remaining gaps are in repeat-rich regions near the centromeres. A tiling path of clones spanning this map and STS maps of chromosomes X and 4 was sequenced to low coverage; the maps and tiling path sequence were used to support and verify the whole-genome sequence assembly, and tiling path BACs were used as templates in sequence finishing.     

A genetic linkage map with 178 SSR and 1901 SNP markers constructed using a RIL population in wheat (Triticum aestivum L.)

The construction of high density genetic linkage map provides a powerful tool to detect and map quantitative trait loci(QTLs) controlling agronomically important traits. In this study, simple sequence repeat(SSR) markers and Illumina 9K i Select single nucleotide polymorphism(SNP) genechip were employed to construct one genetic linkage map of common wheat(Triticum aestivum L.) using 191 recombinant inbred lines(RILs) derived from cross Yu 8679×Jing 411. This map included 1 901 SNP loci and 178 SSR loci, covering 1 659.9 c M and 1 000 marker bins, with an average interval distance of 1.66 c M. A, B and D genomes covered 719.1, 703.5 and 237.3 c M, with an average interval distance of 1.66, 1.45 and 2.9 c M, respectively. Notably, the genetic linkage map covered 20 chromosomes, with the exception of chromosome 5D. Bioinformatics analysis revealed that 1 754(92.27%) of 1 901 mapped SNP loci could be aligned to 1 215 distinct wheat unigenes, among which 1 184(97.4%) were located on o ne single chromosome, and the rest 31(2.6%) were located on 2 to 3 chromosomes. By performing in silico comparison, 214 chromosome deletion bin-mapped expressed sequence tags(ESTs), 1 043 Brachypodium genes and 1 033 rice genes were further added onto the genetic linkage map. This map not only integrated genetic and physical maps, SSR and SNP loci, respectively, but also provided the information of Brachypodium and rice genes corresponding to 1 754 SNP loci. Therefore, it will be a useful tool for comparative genomics analysis, fine mapping of QTL/gene controlling agronomically important traits and marker-assisted selection breeding in wheat.     

小麦RIL群体遗传连锁图谱的构建及其多态性分析

以普通小麦重组近交系(recombinant inbred lines,RIL)‘Q9086×陇鉴19’为作图群体,利用SSR标记构建小麦遗传连锁图谱.结果表明:通过选用2 187对SSR引物筛选出RIL群体双亲表现多态性的引物共405对,多态性频率为18.52%.不同类型SSR标记多态性频率从小到大依次为Xpsp(4.4%)相似文献   

利用水稻BAC克隆对Gm-2和Gm-6在药用野生稻中的FISH定位

 采用栽培稻遗传图第 4连锁群中与抗稻瘿蚊基因 ,Gm- 6和 Gm- 2等位的 RFL P标记 RG2 14和 RZ5 6 9筛选出来的两个 BAC克隆为探针 ,对药用野生稻进行了荧光原位杂交物理定位。两个 BAC克隆的大小分别为 5 0 kb和86 kb,在 Cot- 1DNA封阻的情况下它们均被定位于药用野生稻第 4染色体长臂 ,与着丝粒百分距为 72 .33± 4.40、77.10± 2 .40 ,信号检出率分别为 6 1.2 %和 5 9.5 %。与此同时 ,用 RG2 14和 RZ5 6 9对药用野生稻进行了杂交 ,它们也被定位于药用野生稻第 4染色体长臂 ,与着丝粒百分距分别为 74.18± 2 .6 2和 78.2 3± 2 .31,信号检出率分别为 8.3%和 9.4%。 BAC克隆的 RFL P标记探针杂交位置几乎一致 ,这表明在栽培稻和野生稻中 RFL P标记 RG2 14和RZ5 6 9都在同一 BAC克隆的大插入片段中 ,药用野生稻与抗性基因 Gm- 6和 Gm- 2同源顺序就在第 4染色体信号出现的相应位置。药用野生稻第 4染色体的确定是根据 Jena等 (1994)和本研究的 RFL P的杂交结果进行的。文中讨论了利用栽培稻 BAC克隆对药用野生稻进行原位杂交物理作图的可行性等问题。     

The Physical Location of Gm-2 and Gm-6 in O.officinalis with BAC-FISH Based on Comparative RFLP Map of Wild Rice and Cultivated Rice

Rice BAC library is used widely in rice genome research due to its distinctive advantages over other library systems. In this study, two rice BAC clones closely linked to rice gall midge resistance, Gm-2 and Gm-6, were in situ hybridized to Oryza officinalis chromosomes. They were located on the long arm of chromosome 4 with FL 72.33% and 77.10% respectively and their FL was consistent with the selective marker of rice, RG214 and RZ569. The frequency of signal detection was 61.2% and 59.5% respectively.Our study was based on comparative RFLP map of wild rice, O. officinalis, and cultivated rice, O. sativa.     

PCR-based screening of BAC clones of different chromosomes in Chinese cabbage

In this paper, taking SSR and functional gene sequence as the primers and the plasmid of first- and second-level pools of bacterial artificial chromosome (BAC) library as templates, the PCR method was used for specific clones of different chromosomes in Chinese cabbage. The results showed that the number of positive clones was 1?C11 per primer and the average number of clone was 3.9 by screening 19200 clones of BAC library using 12 pairs of SSR primers from 10 linkage groups individually, which were nearly consistent with about 3.4 times of genome coverage. Positive clones were acquired in chromosome Nos. 2 to 5 and 8 to 10 without screening with the positive clones in chromosome Nos. 1, 6, and 7. In addition, the primer of FLC1 functional gene of chromosome No. 10 was used for PCR screening, and two BAC clones containing FLC1 gene were acquired. Therefore, different specific BAC clones of chromosomes were taken by using SSR primer and functional gene primer. Specific clone screening of chromosomes could provide a probe for identifying the chromosome accurately. Meanwhile, the BAC library screening method was optimized, serving as an effective technical means for quick BAC clone screening.     

The Meiotic Behavior of an Alien Chromosome in Triticum aestivum-Haynaldia villosa Monosomic Addition Lines

By the combination of cytological analysis and using genomic in situ hybridization technique to identify an alien chromosome in wheat-Haynaldia villosa monosomic addition lines, we studied the meiotic behavior of the alien chromosome. The results indicated that the frequency of bivalent pairing was lower than the value expected in PMCs of two monosomic addition lines, the frequency of wheat chromosomes unpairing increased, and the wheat homologous chromosome pairing was interfered with by the added chromosome 6V at metaphase I. The chromosome 6V lagged in 20.3% -29.3% of PMCs, sister chromatids 6V early divided in 29.0% - 34.1% of PMCs, the single chromosome 6V in 18.2% - 26.1% of PMCs went to a pole randomly,the breakage frequency of chromosome 6V was 1.2% - 2.9%. Meanwhile, it was also found that several wheat chromosomes showed earlier division, lagging and breakage in a few PMCs. It revealed that the added chromosome 6V influenced the behavior of wheat chromosomes at anaphase. It was also found that the translocation was produced between 6V and wheat chromosomes in 1.2% of PMCs. It offered evidence for translocation between wheat and Haynaldia villosa 6V chromosomes.     

普通小麦与华山新麦草衍生后代的农艺性状和细胞遗传学研究

对中国春ph2b突变体(CSph2b)与华山新麦草属间杂种F1的衍生后代(BC2和BC1F1)进行了细胞学观察和田间农艺性状调查。其主要结果为:①染色体数目从2n=42-52之间变化,其中2n=45出现频率最高,为20.27%,而2n=51最低,仅有0.66%;②大部分植株有较多单价体;③在染色体数目较多的植株中,能够观察到三价体、四价体、落后染色体、三分孢子体、微核以及染色体桥;④部分植株表现高抗条锈病和赤霉病;⑤各个性状的变异系数都大于亲本的变异系数,分蘖的变异系数最大;⑥2n=42和2n=44的植株细胞学相对稳定,为获得华山新麦草染色体的附加系、代换系或易位系等小麦遗传育种材料奠定了基础。     

黑粒小麦‘冬黑10号’的细胞学和麦谷蛋白组成分析

山西省作为中国优质小麦重要产区,目前已育成6个黑粒小麦品种,然而对这些黑粒小麦的染色体构成、蛋白组分却研究较少。因此,了解其遗传组成对选育黑粒小麦新品种具有指导作用。利用染色体C-分带,减数分裂中期Ⅰ染色体配对构型、基因组原位杂交结合谷蛋白亚基分析,对黑粒小麦‘冬黑10号’的染色体构成进行系统鉴定。结果表明,‘冬黑10号’体细胞染色体数目为2n=42,染色体带型与‘中国春’染色体带型未见明显变化,基因组原位杂交也未检出赖草染色体;其PMC MⅠ染色体平均构型为21Ⅱ,具有良好的细胞学稳定性,与‘中国春’小麦杂交F1的PMC MⅠ染色体构型为21Ⅱ,说明其为六倍体普通小麦。SDS-PAGE分析揭示其高分子量麦谷蛋白亚基组成为Null/7+9/5+10,低分子量麦谷蛋白亚基与对照‘中国春’不同。     

小麦TaGA20ox2基因的克隆及分析

 【目的】克隆小麦中的GA20-氧化酶基因并进行遗传定位,为深入研究小麦中的GA20-氧化酶基因作用机理奠定基础,同时为改良小麦的重要农艺性状提供理论依据。【方法】采用同源克隆结合BAC文库筛选的方法克隆基因,利用中国春缺失体以及国际作图群体对该基因进行遗传定位。【结果】从普通小麦中国春中分离得到TaGA20ox2的gDNA与cDNA序列,利用中国春缺失体材料将TaGA20ox2定位于3A、3B、3D染色体上,序列分析表明分别位于3A、3B、3D染色体上的TaGA20ox2-A1、TaGA20ox2-B1和TaGA20ox2-D1与水稻GA20ox2为直向同源基因,具有GA20ox2家族保守结构域;利用国际作图群体W7984×Opata85将位于3D上的基因TaGA20ox2-3定位于xfba330与xgwm664标记之间。【结论】分离获得小麦中分布于第三同源群的GA20ox2;分子标记结果结合QTL作图信息可以初步推测TaGA20ox2-3在小麦中可能是控制株高的基因。     

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.     

小麦易位系的创造研究进展

小麦作为世界上广泛种植的粮食作物之一,其种质资源日益匮乏,创造小麦外源染色体易位系是拓宽小麦遗传基础的有效途径。创造易位系的方法有利用电离辐射、利用杀配子染色体、通过调节Ph基因的作用诱发部分同源染色体交换、通过染色体错分裂、利用组织培养诱导易位。对这几种创造小麦易位系的方法进行了综述,并作出展望,为小麦育种提供基础。     

小麦和无融合生殖小麦草杂交后代(BC_2F_2)的生物学特性及根尖染色体数目变化研究

对小麦 (Triticumaestivum)和无融合生殖小麦草 (Elymusrectisetus)的杂种后代 (BC2 F2 )生物学特性和根尖染色体数目变化情况进行了的研究。结果表明 ,BC2 F2 的生物学特性处于小麦和小麦草之间 ,但更趋向于小麦 ,这是由于杂交后代经 2次回交所致。根尖染色体检查发现 ,BC2 F2 的体细胞染色体数目存在 1个很大的变异范围 ,从 2 9条到 6 9条不等 ,但大部分细胞内的染色体数目是 42条。杂种后代的体细胞染色体数目的变化一方面与远缘杂交有关 ,另一方面可能与无融合生殖具有一定的联系     

普通小麦-百萨偃麦草染色体易位的选育与鉴定

 利用染色体C 分带、基因组原位杂交和花粉母细胞减数分裂分析的方法 ,从普通小麦中国春与中国春 百萨偃麦草双倍体回交BC1F5 代中选育出 5份纯合易位新种质 ,分别为 :含 1对易位染色体并添加 1对百萨偃麦草完整染色体的Tj0 1和Tj0 2 ;添加 1对易位染色体的Tj0 3 ;含 1对易位染色体并添加 1对百萨偃麦草某一染色体臂端着丝粒染色体的Tj0 4和含 1对易位染色体并添加 2对百萨偃麦草完整染色体的Tj0 5。这些易位染色体的易位断点均不在着丝粒处 ,能稳定传递 ,且所涉及的植株生长和结实均正常。推测在该回交后代中小麦与百萨偃麦草染色体之间发生了自发的部分同源重组 ,这将有利于百萨偃麦草优异基因向普通小麦的转移与利用     

Development and Identification of Triticum aestivum L.-Thinopyrum bessarabicum L(o)ve Chromosome Translocations

With ass7istance of chromosome C-banding and genomic in situ hybridization(GISH)combined with meiotic analysis,five germplasms with homozygous wheat-Th. Bessarabicum chromosome translocations were developed and identified among BC1F5 progenies of the cross between T. Aestivum cv. Chinese Spring and Chinese Spring-Th. Bessarabicum amphiploid. These lines included Tj01 and Tj02(2n=44)containing a pair of wheat-Th. Bessarabicum translocation chromosomes besides a pair of added Th. Bessarabicum chromosomes,Tj03(2n=44)with a pair of added interspecific translocation chromosomes,Tj04(2n=44)containing a pair of interspecific translocation chromosomes besides an added pair of Th. Bessarabicum chromosome arms and Tj05(2n=46)containing a pair of interspecific translocation chromosomes besides two pairs of added intact alien chromosomes. The breakpoints of all the translocations were found to be not around centromere. Meanwhile,all the lines showed normal plant growth,development and fertility,while the translocation chromosomes transmitted regularly. The obtained translocations might be of use for transferring elite genes from Th. Bessarabicum into wheat.     

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。     

小麦品种偃展1号与品系早穗30重组自交系群体遗传连锁图谱构建及重要农艺性状的QTL分析

【目的】构建重组自交系(recombinant inbred line,RIL)群体及其遗传连锁图谱,对小麦重要农艺性状进行数量性状位点(quantitative trait locus,QTL)分析,为发现小麦新基因与分子标记辅助育种奠定基础。【方法】配制普通小麦品种(系)早穗30和偃展1号的杂交组合,通过一粒传的方法培育重组自交系群体;利用SSR(simple sequence repeat)标记、DarT(diversity arrays technology)标记、ISBP(insertion site-basedpolymorphism)标记以及抽穗期和株高的功能标记绘制其遗传连锁图谱并通过复合区间作图法(Compositeinterval mapping,CIM)对多个环境下的抽穗期、株高、千粒重、穗粒数、每穗小穗数、穗长等农艺性状进行QTL定位分析。【结果】培育出由219个F7家系组成的重组自交系群体;构建了含481个分子标记的遗传连锁图谱;检测出分布在12条染色体上的26个与重要农艺性状相关的QTL,其中9个QTL能够在至少2个环境下重复;研究还发现了3个QTL聚集的"QTL簇",其中4D染色体上的矮秆基因Rht2所在区段控制株高与千粒重,5D染色体上的Vrn-D1-WMS212区间控制抽穗期、穗粒数与每穗小穗数,7B染色体上wPt4230-wPt4814区段控制抽穗期、穗粒数、株高与穗长。【结论】构建的小麦遗传作图群体可成功地用于重要农艺性状分析;矮秆基因Rht2与春化基因Vrn-D12个发育相关基因均与多个重要农艺性状有关;在7B上可能存在与发育相关的重要新基因。     

盐胁迫下调控小麦苗期性状的QTL分析

【目的】定位盐胁迫下调控小麦苗期性状的QTL位点,为分子标记辅助选择小麦耐盐性状提供基因位点和连锁标记。【方法】以小偃54×京411重组自交系群体为材料,在盐胁迫条件下检测调控小麦苗期MRL、     RDW、SDW和TDW及其相对性状的QTL位点。【结果】共检测到调控小麦苗期4个性状及其相对性状的25个QTL位点,分布在1A、2A、2D、3A、4A、4B、5B、5D、6B、7A和7B共11条染色体上,贡献率在4.4%-25.5%。其中有15个QTL位点成簇分布于3A、4A、4B、5B、5D染色体的5个遗传区间,其余10个QTL位点各自分布于不同的染色体区段。检测到的5个贡献率大于10%的位点分别位于3A染色体的Xgwm497.1-Xcfa2193和4A染色体的Xbarc78-Xgwm350.1。【结论】多数调控小麦耐盐性的QTL位点成簇分布于3A、4A、4B、5B和5D染色体上,3A染色体的Xgwm497.1-Xcfa2193和4A染色体的Xbarc78-Xgwm350.1携带所有5个贡献率在10%以上的QTL位点。     

芥菜型油菜A9染色体黄籽基因区域BAC重叠群的构建

A9染色体是芸薹属植物A基因组最长的染色体(序列约37 Mb),具有控制种子大小、种皮颜色、硫苷合成、油脂合成等重要性状的基因。利用芥菜型油菜A9染色体黄籽性状共分离标记A9-88和A9-32对已经构建的ZBjH BAC文库进行PCR步移筛选,共筛选出BAC 752个,测序395个,得到BAC末端序列674条,利用BAC末端序列设计引物533对,此外利用白菜已经公布的序列设计SSR引物38对、STS引物15对,构建了芥菜型油菜黄籽基因区域估算长约2.2 Mb的BAC重叠群。     

普通小麦5DL染色体分选及染色体BIBAC文库构建技术体系优化

 【目的】优化小麦染色体流式分选及特定染色体BAC文库构建技术体系,为加速小麦基因组学研究提供技术支撑。【方法】采用双阻断法对中国春5DL端体根尖分生组织进行细胞周期同步化处理,结合机械匀浆法制备染色体悬浮液进行流式核型分析,分选染色体5DL及PCR检测,对小麦染色体分选技术体系进行优化。对复合峰分选产物Bam HI酶切制备高分子量（high molecular weight,HMW）DNA,按10﹕1质量比与双元细菌人工染色体（binary-bacterial artificial chromosome,BIBAC）连接,电激转化感受态细胞DH10B,随机挑取100个重组子,经NotI完全酶切后脉冲电泳检测插入片段大小,优化小麦染色体BAC文库构建的技术体系。【结果】经1.25 mmol•L-1 羟基脲（hydroxyurea,HU）18 h,恢复6 h和1 µmol•L-1 氟乐灵（trifluralin）2.5 h的顺序处理的小麦根尖分生组织,细胞有丝分裂指数可超过60%。流式核型分析表明,5DL端体的流式核型图由清晰的三个复合峰和两个单峰组成。PCR鉴定表明,其中一单峰确由5DL染色体组成。部分酶切复合峰染色体3～10 min,可以获得50～300 kb 的HMW DNA。连接产物检测显示平均插入片段可达100 kb左右,根据荧光通道值,估算5DL端体的大小约为482 Mb,构建库容为19 000个克隆的5DL端体特异文库即可达到约4倍的覆盖率。【结论】上述两项优化技术适用于小麦染色体的分选及染色体特异文库的构建。