大豆EST-SNP的挖掘、鉴定及其CAPS标记的开发

采用生物信息学方法将大豆EST序列联配到大豆基因组序列上,挖掘到大豆EST-SNP位点537个。对其靶向基因进行功能注释分析,发现他们主要参与亚细胞定位、蛋白质结合与催化以及代谢等与大豆重要农艺性状形成相关的生物过程。同时开发了简便易行的SNP检测方法,利用EMBOSS软件筛选导致酶切位点改变的EST-SNP,分别以大豆绥农14、合丰25、Acher、Evans、Peking、PI209332、固新野生大豆、科丰1号、南农1138-2的DNA及其混合的DNA为模板,设计引物进行PCR扩增,发现44个PCR产物中有36个测序峰图在预期的EST-SNP位点表现出多态性。酶切分析发现26个PCR产物具有酶切多态性,可以作为CAPS标记。结果表明该EST-SNP挖掘体系及其CAPS标记转化系统具有高效率、低成本等优点,有利于促进大豆的遗传育种研究。     

油松猝倒病病原菌鉴定及致病性测定

[目的]油松苗期发生的猝倒病是油松育苗工作的极大障碍,对该病害病原菌的正确鉴定是研究其致病机理、开展防治工作等的基础。[方法]对1株引起油松苗猝倒病的病原真菌通过形态学、培养性状、致病性及rDNA ITS序列测定来鉴定该病原菌。[结果]在PDA培养基上,该病原菌幼嫩菌丝无色,但在生长过程中逐渐变为褐色,部分菌丝膨大成念珠状;菌丝细胞多核;菌丝平均直径6~10μm,分支发生点附近缢缩并形成一隔膜,分支与再分支呈直角;不产生无性孢子,也不产生菌核;能引起油松苗猝倒病;其rDNAITS序列与瓜亡革菌（Thanatephorus cucumeris）的2个菌株的相似性最大,均为97%。[结论]该病原菌被鉴定为立枯丝核菌（Rhizoctonia solaniKhn）。该研究可为后续试验的进行奠定基础。     

斯卑尔脱小麦α-醇溶蛋白基因克隆与序列分析

【目的】进一步了解斯卑尔脱小麦（Triticum spelta L.）α-醇溶蛋白基因序列信息。【方法】根据已知的普通小麦α-醇溶蛋白基因序列设计引物,采用PCR方法,克隆基因并进行序列分析。【结果】从NGB5149中克隆得到两个α-醇溶蛋白基因序列Gli-Spelt-1和Gli-Spelt-2（GenBank登录号分别为DQ234066和DQ234067）。它们具有α-醇溶蛋白基因的典型结构特征,但Gli-Spelt-1是一个假基因。Spelt-Gli-2编码区全长849 bp,编码263个氨基酸。【结轮】氨基酸序列比较显示,Gli-Spelt-1和Gli-Spelt-2与已报道的α-醇溶蛋白序列有较高的一致性。     

新疆加工番茄上番茄花叶病毒的分子鉴定

从表现花叶、畸形的加工番茄病株上获得分离物xj124,用1 对ToMV CP 基因引物进行 RT-PCR ,扩增得到了689 bp 的特异性核苷酸片段.将PCR产物插入到克隆载体pMD1 8-T中再转化到大肠杆菌TOP10.经限制酶酶切鉴定,重组克隆中含有与PCR 产物大小相同的689 bp 的插入片段.cDNA全序列分析表明,所扩增出的689 bp ToMV CP基因,含1个480 bp开放阅读框架( ORF),编码160氨基酸组成的蛋白,所克隆的基因包含完整的ToMV CP基因.所测得的xj124分离物与国内外已报道的部分ToMV CP基因序列比较,核苷酸同源性高达84.4;～99.8;,氨基酸同源性高达98.7;～100;.因此将该分离物鉴定为番茄病毒病花叶病毒.     

利用rbcL基因序列比对分析广西优质红锥种质资源的遗传多样性

通过PCR扩增得到10个红锥品系的rbcL基因全序列,均为1.513 kb.核苷酸序列比对结果表明,在256-1 455 bp区间10个红锥品系共有16处碱基发生变异,变异率为1%,其中在306 bp和634 bp处10个红锥品系较同属对照castanopsis lucida均发生相同的变异,可以看作属内进化信息碱基.对表达的氨基酸序列进行比对分析,发现16处碱基变异共引起8处氨基酸序列变异.     

不同症状矮牵牛植株植原体16SrDNA 片段的克隆及序列分析

应用植原体16SrRNA基因通用引物,分别对表现矮化和扁茎症状的矮牵牛植株进行巢式PCR检测,均得到约1．2kb的特异片段。将此特异片段与pGEM-T Easy载体连接并转化到大肠杆菌JM109感受态细胞中,通过PCR鉴定,序列测定及同源性比较分析,结果表明：这2个植原体株系16SrDNA片段G C含量分别为47．1％和47．0％,具有其他柔膜菌纲原核生物16SrRNA基因碱基组成特征;与16Sr组中的代表株系(西方翠菊黄化,SAY)同源率达到最高,分别为99．1和98．6％,故认为这2个株系为该组成员之一。     

黄槐丛枝病植原体的检测及鉴定

 应用植原体16S rRNA基因通用引物,对自然表现丛枝的黄槐植株进行巢式PCR检测,得到约1.2 kb的特异片段,证明此植株中存在植原体.将此特异片段与pGEM-T Easy载体连接并转化到大肠杆菌JM109感受态细胞中,通过PCR鉴定、序列测定及同源性比较分析,结果表明此植原体株系(STWB)16S rDNA片段G+C含量为45.8%,与榆树黄化植原体组(Elm yellows group,16SrV group)中的各株系最高同源率可达99.4%,而与其它组中的株系明显低于97.0%,故认为该植原体株系为榆树黄化植原体组中的成员之一.     

Inter-Alu PCR like genomic profiling in banana

Alu sequences are a major repetitive element of the primate genome. To date Alu sequences have seldom been reported in plant genomes. We report here an inter-Alu PCR like genomic profiling in banana using a single primer designed from a human Alu sequence. The different Musa species involved in the complex genome of banana cultivars can thus be discriminated. The use of this technique for monitoring germplasm or genotyping cultivars is discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genome-wide development of interspecific microsatellite markers for Saccharum officinarum and Saccharum spontaneum

Sugarcane has a large, complex, polyploid genome that has hindered the progress of genomic research and molecular marker-assisted selection. The user-friendly SSR markers have attracted considerable attention owing to their ideal genetic attributes. However, these markers were not characterized and developed at the genome-wide scale due to the previously lacking high-quality chromosome-level assembled sugarcane genomes. In this present study, 744305 and 361638 candidate SSRs were identified from the genomes of S. officinarum and S. spontaneum, respectively. We verified the reliability of the predicted SSRs by using 1200 interspecific SSR primer pairs to detect polymorphisms among 11 representative accessions of Saccharum, including S. spontaneum, S. officinarum, S. robustum, and modern sugarcane hybrid. The results showed that 660 SSR markers displayed interspecific polymorphisms among these accessions. Furthermore, 100 SSRs were randomly selected to detect the genetic diversity for 39 representative Saccharum accessions. A total of 320 alleles were generated using 100 polymorphic primers, with each marker ranging from two to seven alleles. The genetic diversity analysis revealed that these accessions were distributed in four main groups, including group I (14 S. spontaneum accessions), group II (two S. officinarum accessions), group III (18 modern sugarcane hybrid accessions), and group IV (five S. robustum accessions). Experimental verification supported the reliability of the SSR markers based on genome-wide predictions. The development of a large number of SSR markers based on wet experiments is valuable for genetic studies, including genetic linkage maps, comparative genome analysis, genome-wide association studies, and marker-assisted selection in Saccharum.     

香蕉枯萎病菌4个生理小种内切多聚半乳糖醛酸酶基因的比较

本研究通过基因克隆和测序方法，对来自国内外Foc 的1号和4号小种的6个菌株endo-PG基因同源性、与其他真菌的亲缘关系以及endo-PG基因和氨基酸序列进行了分析，为香蕉枯萎病菌致病机制的研究奠定基础。结果表明，同一小种的endo-PG序列同源性高、亲缘关系近，不同小种的同源性较低、亲缘关系较远；总体来说，4号小种比1号小种同源性更高、亲缘关系更近。在全基因序列分析中，并没有发现明显的基因差异可以作为区分2个小种致病性差异的依据。根据预测的CDS序列翻译成氨基酸序列比对，发现6个菌株的endo-PG基因编码的氨基酸变异并没有引起相关保守结构域的变化，推测6个菌株的致病性可能与endo-PG基因的调控相关。