TAI系列Ⅰ中的冰草染色体与小麦部分同源关系的同工酶研究

本文分析了TAI系列Ⅰ及其亲本的胚乳过氧化物酶(CPXE)、苹果酸脱氢酶(MDH)、醇脱氢酶(ADH)、酸性磷酸酶(ACPH)和碱性磷酸酶(PHE)的酶谱表型。把冰草同工酶结构基因Mdh-Ag~t2定位到TAI-13、基因Cpxe-Ag~2定位到TAI-14、基因Adh-Ag~t1、Acph-Ag~t2和Phe-Ag~t2定位到TAI-16中的冰草染色体上。根据这些基因定位和我们以前的研究工作,结合植株表型分析,推测TAI-11、TAI-12、TAI-13、TAI-14、TAI-15、TAI-16和TAI-17中的冰草染色体分别与小麦第2、3、1、7、6、4和5同祖群有一定部分同源关系。     

TAI系列Ⅱ天兰冰草染色体上的酯酶同工酶基因定位

本文分析了小冰麦异附加系列Ⅱ,八倍体小冰麦,普通小麦和天兰冰草的叶片酯酶（ESTL）,胚乳酯酶（ESTE）,根酯酶（ESTR）和胚芽鞘酯酶（ESTG）的酶谱表型。把冰草同工酶基因Estl-Ag1,Este-Agi1,Estr-Agij,Estc-Agi1,Estl-Agi2和Este-Agi2定位到异附加系TAI－23,把基因Estr-Agi4和Estc-Agi4定位到TAI－24中的冰草染色体上,根据这些基因定位,结合某些植株形态学特征,推测TAI－23和TAI－24中的冰草染色体分别与小麦第3和6同祖群有一定的部分同源关系。     

小冰麦异附加系中天兰冰草染色体的变异

通过对两套14种小冰麦异附加系的体细胞染色体观察,发现3个异附加系各有1对染色体发生了显著变异。其中TAl-14t有1对端着丝点染色体,TAl-22s和TAl-27s各有1对近中着丝点的小染色体。通过组胞遗传学分析,从3个方面证明了发生变异的染色体均来自天兰冰草。此外还发现在其中的两个异附加系(TAI-14t和TAl-27s)中可能产生了自发易位。     

TAI系列I中的冰草染色体与小麦部分同源关系的同工酶研究

本文分析了ＴＡＩ系列Ｉ及其亲本的胚乳氧化物酶,苹果酸脱氢酶,醇脱氢酶,酸性磷酸酶和碱性磷酸酶的酶谱表型。把冰草同工酶结构基因Ｍｄｈ－Ａｇ＾ｉ２定位一ＴＡＩ－１３,基因Ｃｐｘｅ－Ａｇ＾ｉ２定位到ＴＡＩ－１４,基因Ａｄｈ－ＡＧ＾Ｉ１,Ａｃｐｈ－Ａｇ＾ｉ２和Ｐｈｅ－Ａｇ＾ｉ２定位到ＴＡＩ－１６中的冰草染色体上。根据这些基因定位和我们以前的研究工作,结合植株表型分析,推测ＴＡＩ－１１,ＴＡＩ－１２,ＴＡＩ     

天兰冰草染色体对小冰麦外植体再生能力的影响及基因的染色体定位

本文研究了天兰冰草（Ａｇｒｏｐｙｒｏｎｉｎｔｅｒｍｅｄｉｕｍ２ｎ＝４２）染色体导入小麦后对小冰麦外植体再生能力的影响,从而分析调控再生能力的基因所在天兰冰草染色体的定位。对异源八倍体小冰麦的研究表明,与天兰冰草强再生能力有关的基因主要分布在附加到中２的冰草染色体组上;对７个异附加系小冰麦的研究进一步表明,冰草强再生能力的性状由多基因调控,这些基因主要分布在附加到ＴＡＩ－１１、ＴＡＩ－１２、ＴＡＩ－１３的冰草染色体上。此外,小冰麦杂种Ｆ１的研究表明,在组织培养中同样能够表现出杂种优势。     

小麦新种质4844中外源P染色质的GISH与SSR分析

采用基因组原位杂交(GISH)检测和染色体组成分析方法,对大穗多花小麦新种质4844后代的15个株系进行遗传分析。结果发现,4844-12是1个稳定的异附加系,4844-2和4844-8是稳定的异代换系;对异代换系进行SSR分析表明,代换系中小麦的6D染色体被1对P染色体代换,说明这对冰草染色体与小麦6D染色体有部分同源关系,由此确定4844中的冰草染色体为6P;同时筛选出冰草6P染色体的4个SSR标记。     

家蚕性连锁平衡致死系致死基因的SSR定位

家蚕性连锁平衡致死系(S-14)雄蚕的两条Z染色体分别携带有一个非等位、紧密连锁的隐性胚胎期致死基因l1(lethal gene1)和l2(lethal gene2)。两个致死基因的致死时期分别是转青期和G2期。将S-14品系的雄蚕和家蚕P50品系的野生型雌蚕杂交,F1代雄蚕和P50品系雌蚕回交,即P50×(P50×S14)。回交后代雌蛾根据父本(F1代雄蚕)携带l1或l2基因分成两类BC1-l1和BC1-l2,分别用来做l1和l2基因定位。利用公布的家蚕全基因组序列筛选l1基因和l2基因所在Z染色体与P50品系Z染色体间的差异SSR标记,分别获得16个和18个差异性SSR标记,用差异性标记检测BC1-l1和BC1-l2,最终将l1基因定位在Z染色体物理图谱中的19.79Mb位点到染色体末端约2.60Mb范围内,将l2基因定位在Z染色体物理图谱的17.86Mb位点到18.55Mb位点约0.69Mb范围内。     

小冰麦异附加系TAI系列中异源麦谷蛋白基因位点的生化与分子生物学鉴定

小冰麦异附加系TAI系列的每一个材料中分别附加了1对来自中问偃麦草的染色体,附加染色体很容易丢失,使得失去附加染色体的小麦可以作为异附加系的对照材料。通过分析TAI系列异附加系及各自对照材料的高分子量麦谷蛋白亚基组成与低分子量麦谷蛋白基因的PCR图谱,鉴定出异附加系TAI-13和TAI-25中具有编码中问偃麦草麦谷蛋白的基因位点,附加的中间偃麦草染色体属于第一同源群。异附加系TAI-11中附加的中间偃麦草染色体只具有低分子量麦谷蛋白基因位点。     

小麦—冰草T7PL·7AL罗伯逊易位系的分子细胞学鉴定

冰草是小麦遗传改良的重要野生近缘植物之一,有目标的导入冰草外源优异基因是拓宽小麦遗传基础的有效途径。前期研究表明:小麦-冰草衍生系II-23(2n=38W+6P)由19对小麦染色体(缺少4B和7A)和3对冰草染色体(2P、4P和7P)组成。本研究报道从II-23的回交后代中分离鉴定出1个自发易位系7-20。基因组原位杂交(GISH)鉴定表明7-20是一个整臂易位系;经非变性荧光原位杂交(ND-FISH)检测发现,小麦的7A染色体发生易位;进一步利用小麦7A染色体特异SSR标记以及冰草7P染色体特异STS标记对7-20易位系中的外源易位片段大小以及易位染色体的组成进行鉴定,确定7-20为T7PL·7AL罗伯逊易位系(Robertsonian translocation line)。对该易位系与小麦品种Fukuhokomugi构建的BC1F2和BC2F1世代分离群体进行田间农艺性状考察,发现该易位系阳性株系和阴性株系在有效分蘖数和千粒重性状上无显著差异,在株高上表现为阳性材料显著低于阴性材料,但同时出现穗粒数下降的现象。总之,本研究表明易位系7-20为T7PL·7AL罗伯逊易位,该创新材料不仅为后续利用断裂—融合机制创制出更多的补偿易位材料提供了理论依据,而且也为今后向小麦中转移冰草优异基因提供了重要的中间桥梁材料。     

用基因芯片筛选高转移卵巢癌细胞系转移相关基因

用标准化的Affymetrix公司生产U133A基因芯片技术研究高(H)转移卵巢癌细胞株(HO-8910PM)和正常卵巢上皮(C)基因表达谱差异,筛选与卵巢癌转移相关的基因及其在染色体的定位和功能。结果发现高转移卵巢癌细胞株和正常卵巢上皮比较表达差异8倍以上共有1,237个基因,其中表达上调(信号比的对数值SLR≥3)有597个,表达下调(SLR≤-3)有640个。从表达差异的基因在染色体定位分析,发现除1个基因未知其定位外,其余所有差异表达基因散在分布在各条染色体上,但以1号染色体最多,有115个(9.3%)。其次是2号染色体有94个(7.6%),第三是12号染色体有88个(7.1%)。第四是11号染色体有76个(6.1%)。第五是X染色体有71个(5.7%)。第6是17号染色体有69个(5.6%)。而差异表达的基因发生在染色体短臂(q)上有805个(占65.1%),在13,14,15,21和22号仅发现在q上有差异表达基因。从表达差异的基因分子功能分类看,属于酶和酶调控子基因最多(306个,占24.7%),其次是核酸结合基因(144个,占11.6%)。第三类是信号传导基因(137个,占11.1%)。第四类是蛋白结合基因(116个,占9.4%)。以上4大类共占基因总数56.8%。还有功能未知的基因有207个,占16.7%。结论:高转移卵巢癌细胞株差异表达基因散在分布在各条染色体上,但以1、2、12、11、17和X染色体差异表达基因居多,肿瘤的转移是多基因共同作用的结果。4大类(酶和酶调控子活性、核酸结合活性、信号传导活性、蛋白结合活性)差异表达基因是我们今后研究卵巢癌转移相关的重要基因。     

Variation of the Wheatgrass Chromosomes in Wheat-wheatgrass Disomic Addition Line TAI 14 Revealed by Fluorescence in Situ Hybridization

The karyotype of the primary wheat-wheatgrass disomic addition line TAI-14 was 2n = 44 in which all of the chromosomes were metacentric and submetacentric. However, in the progeny of the TAI-14, a pair of telocentric chromosomes were observed. In order to clarify whether the telocentric chromosomes were of common wheat or of wheatgrass, the fluorescence in situ hybridization (FISH) technique was employed. It was revealed that the wheat chromosomes exhibited red fluorescence while the ditelocentric chromosomes green fluorescence. Therefore, the primary TAI-14 was conversed from the disomic addition line into the ditelocentrie addition line. The possible explanation for such a variation and the potential significance of the ditelocentric addition line were discussed briefly.     

Variation of wheatgrass chromosomes in wheat-wheatgrass alien addition line "TAI-27"revealed by fluorescence in situ hybridization (FISH)

The Karyotyp of the primary wheat-whastgrass alien addition line TAI-27 was 2n = 44 in which all d the chromosomes were metacentric and subrmetacentric. However, in the progeny of TAI-27 a pair of chromosomes had become small chromosomea in the two morphologically different plants. Fluorescence in situ hybridizstionm (FISH) technique was used to analyze the two different plants. The observations indicate that a pair of small chromosomes in one varietion line are from wheatgrass. In another variation line, a pair of small chromosomes are also from whest-grass, while another pair of wheatgrass chromosomes have substituted the wheat chromosomes. TAI-27 and its variant lines showed a high level of resistance to barley yellow dwarf virus (BYDV). The pessible explanation for such a variation and the potential use of the variant lines were discussed briefly.     

Variation of wheatgrass chromosomes in wheat-wheatgrass alien addition line “TAI-27” revealed by fluorescencein situ hybridization (FISH)

The Karyotyp of the primary wheat-whastgrass alien addition line TAI-27 was 2n = 44 in which all d the chromosomes were metacentric and subrmetacentric. However, in the progeny of TAI-27 a pair of chromosomes had become small chromosomea in the two morphologically different plants. Fluorescence in situ hybridizstionm (FISH) technique was used to analyze the two different plants. The observations indicate that a pair of small chromosomes in one varietion line are from wheatgrass. In another variation line, a pair of small chromosomes are also from whest-grass, while another pair of wheatgrass chromosomes have substituted the wheat chromosomes. TAI-27 and its variant lines showed a high level of resistance to barley yellow dwarf virus (BYDV). The pessible explanation for such a variation and the potential use of the variant lines were discussed briefly.     

中间偃麦草染色体组构成的同工酶研究

应用聚丙烯酰胺凝胶电泳,研究了带有不同染色体组的各种小麦和中间偃麦草的酯酶、苹果酸脱氢酶、酸性或碱性磷酸酶同工酶的酶谱。通过对各酶谱与染色体组的对比分析表明,中间偃麦草不含与小麦B组或D组同源的染色体,而可能含有两组分别与小麦A组和提莫菲维小麦G组有些同源性的染色体。中间偃麦草的染色体组构成可用E_(A1)E_(A2)N_G或E_(G1)E_(G2)N_A表示。     

Screening and analysis of differentially expressed genes from an alien addition line of wheat Thinopyrum intermedium induced by barley yellow dwarf virus infection.

The alien addition line TAI-27 contains a pair of chromosomes of Thinopyrum intermedium that carry resistance against barley yellow dwarf virus (BYDV). A subtractive library was constructed using the leaves of TAI-27, which were infected by Schizaphis graminum carrying the GAV strain of BYDV, and the control at the three-leaf stage. Nine differentially expressed genes were identified from 100 randomly picked clones and sequenced. Two of the nine clones were highly homologous with known genes. Of the remaining seven cDNA clones, five clones matched with known expressed sequence tag (EST) sequences from wheat and (or) barley whereas the other two clones were unknown. Five of the nine differentially expressed sequences (WTJ9, WTJ11, WTJ15, WTJ19, and WTJ32) were highly homologous (identities >94%) with ESTs from wheat or barley challenged with pathogens. These five sequences and another one (WTJ18) were also highly homologous (identities >86%) with abiotic stress induced ESTs in wheat or barley. Reverse Northern hybridization showed that seven of the nine differentially expressed cDNA sequences hybridized with cDNA of T. intermedium infected by BYDV. Three of these also hybridized with cDNA of line 3B-2 (a parent of TAI-27) infected by BYDV. The alien chromosome in TAI-27 was microdissected. The second round linker adaptor mediated PCR products of the alien chromosomal DNA were labeled with digoxygenin and used as the probe to hybridize with the nine differentially expressed genes. The analysis showed that seven differentially expressed genes were homologous with the alien chromosome of TAI-27. These seven differentially expressed sequences could be used as ESTs of the alien chromosome of TAI-27. This research laid the foundation for screening and cloning of new specific functional genes conferring resistance to BYDV and probably other pathogens.     

Cloning, sequencing, and disruption of the gene encoding sterol C-14 reductase in Saccharomyces cerevisiae.

A sterol C-14 reductase (erg24-1) mutant of Saccharomyces cerevisiae was selected in a fen1, fen2, suppressor background on the basis of nystatin resistance and ignosterol (ergosta-8,14-dienol) production. The erg24-1 allele segregated genetically as a single, recessive gene. The wild-type ERG24 gene was cloned by complementation onto a 12-kb fragment from a yeast genomic library, and subsequently subcloned onto a 2.4-kb fragment. This was sequenced and found to contain an open reading frame of 1,314 bp, predicting a polypeptide of 438 amino acids (M(r) 50,612). A 1,088-bp internal region of the ERG24 gene was excised, replaced with a LEU2 gene, and integrated into the chromosome of the parental strain, FP13D (fen1, fen2) by gene replacement. The ERG24 null mutant produced ergosta-8,14-dienol as the major sterol, indicating that the delta 8-7 isomerase, delta 5-desaturase and the delta 22-desaturase were inactive on sterols with the C14 = 15 double bond.     

An organic-solvent-tolerant esterase from thermophilic Bacillus licheniformis S-86

A thermophile, halotolerant and organic-solvent-tolerant esterase producer Bacillus sp. S-86 strain previously isolated was found to belong to Bacillus licheniformis species through morphological, biochemical, 16S rRNA gene sequence analyses and rDNA intergenic spacers amplification (ITS-PCR). The strain can grow at 55 degrees C in presence of C2-C7 alkanols (log P=-0.86 to 2.39), and NaCl concentrations up to 15% (w/v). This bacterium showed optimal growth and esterase production at 50 degrees C. Two different molecular weight esterase activities were detected in zymographic assays. PMSF inhibited type I esterase activity, showing no inhibitory effect on type II esterase activity. B. licheniformis S-86 was able to grow in presence of hydroxylic organic-solvents like propan-2-ol, butan-1-ol and 3-methylbutan-1-ol. At a sub-lethal concentration of these solvents (392 mmoll(-1) propan-2-ol; 99 mmol l(-1) butan-1-ol, 37 mmol l(-1) 3-methylbutan-1-ol), adequate to produce 50% cell growth inhibition at 50 degrees C, an increment between 1.9 and 2.3 times was observed in type I esterase production, and between 2.2 and 3.1 times in type II esterase production.