油菜叶片衰老相关基因的分离：ATP硫化酶cDNA的克隆和序列分析

通过筛选缩减ｃＤＮＡ文库中在油菜叶片衰老阶段表达而在绿叶中不表达的ｃＤＮＡ克隆，分离了与油菜叶片衰老有关的基因ＬＳＣ６６。ＤＮＡ序列测定和分析结果表明，ＬＳＣ６６的全长可读框架由１３８０个碱基组成，编码４６０个氨基酸，它的核昔酸序列与拟南芥菜ＡＴＰ硫化酶基因有８６．６７％的同源性，其推导的氨基酸序列与拟南芥菜ＡＴＰ硫化酶的氨基酸序列有９１．７４％的同源性。对ＬＳＣ６６基因产物在植物叶片衰老过程中的作用进行了讨论。     

杂交稻对白叶枯病菌主基因抗性的配合力分析

以８种不同胞质类型的不育系及其保持系与９个恢复系配制的７２个不育系杂种及保持系杂种为材料，研究了杂交稻对白叶枯病菌主基因抗性的配合力。结果表明，杂交稻的抗性以主基因抗性为主，但同时也受双亲中与抗性有关的其他加性、显性及上位性基因效应的影响。说明杂交稻的抗性强弱取决于双亲抗性基因互补和累加程度高低。     

Functional analysis of the venom allergen-like protein gene from pine wood nematode Bursaphelenchus xylophilus using a baculovirus expression system

The pine wood nematode, Bursaphelenchus xylophilus, infects pine trees, leading to fatal pine wilt disease. Here, recombinant venom allergen-like protein (VAP) was obtained by expressing Bx-vap-1 in insect cells. Three-year-old Pinus massoniana were inoculated with recombinant VAP, simulating B. xylophilus esophageal gland secretions. Recombinant VAP up-regulated α-pinene synthase gene expression, the trees showed disease symptoms 15 d after inoculation and the xylem pith revealed brown tissue discoloration, indicating that recombinant VAP could damage P. massoniana cells. Recombinant VAP did not, however, lead to cavitation, indicating that the VAP secreted from B. xylophilus acts as a defense response elicitor.     

Pseudomonas putida BP25 alters root phenotype and triggers salicylic acid signaling as a feedback loop in regulating endophytic colonization in Arabidopsis thaliana

Endophytic Pseudomonas putida BP25 (PpBP25) triggered density dependent alterations on Arabidopsis thaliana Col-0 growth. Endogenous colonization of PpBP25 was found regulated within Arabidopsis that caused induction and repression of 131 and 74 plant genes, respectively. Induced genes like WRKY33, AtRLP19, ATL2, ATEXO70B2, pEARLI, RPS2, CBP60G, PLA2, CRK18, ATFBS1, DREB2A, TIR, RAP2.4, and MOS1 were components of defense and salicylic acid (SA) signaling. Development associated genes were found significantly repressed. Biased activation of phytohormone signaling with their associated fitness costs on plant growth was observed. The data suggests that PpBP25 colonization triggered expression of defense genes that restricted its own population in a feedback loop besides causing altered root phenotype.     

柑橘黄化脉明病毒的TGB基因生物信息学分析及亚细胞定位

柑橘黄化脉明病毒(Citrus yellow vein clearing virus,CYVCV)是一种正义单链RNA病毒。其基因组含有6个开放阅读框(ORF),其中ORF2、ORF3和ORF4组成了三基因连锁结构(triple gene block,TGB)。以感染CYVCV的尤力克柠檬[Citrus limon(L.)Burm.f.]植株的总RNA为模板,扩增和克隆了CYVCV的TGB基因,并开展了其编码蛋白的理化性质和分子特性等生物信息学分析;进而通过In-Fusion~?技术构建了TGB各基因的亚细胞定位载体,经农杆菌介导转化洋葱表皮细胞并在荧光显微镜下进行亚细胞定位观察。生物信息学分析结果表明,CYVCV-TGB具有与Potexvirus属病毒TGB相似的理化性质和分子特性,可能参与病毒在寄主中的运动且需要外壳蛋白的参与。亚细胞定位结果显示,TGBp1定位于细胞壁(膜)上,TGBp2和TGBp3主要定位于细胞壁(膜),少量呈星点状定位于细胞内。研究结果为揭示CYVCV在寄主中的运动机理奠定了基础。     

乳腺特异表达载体缺失片段的鉴定与修复

通过对pBC1中的乳腺特异表达启动子成分进行分析,鉴定了位于β酪蛋白启动子上游两个EcoRⅠ位点(-89~-94和-120~-125)之间的缺失片段,31bp的缺失使得该载体中的一个乳控盒元件(-112~-141)和一个乳腺因子识别序列(-92~-102)遭到破坏,并使得位于TATA框上游的所有启动子成分发生了位置改变。通过与山羊β酪蛋白启动子序列进行比较,设计并合成了缺失片段,经一系列的酶切和连接处理,使得该缺失得到了修复,修复后的载体具有所有乳蛋白表达需要的启动子调控原件。本试验结果为利用修复后的乳腺特异高效表达载体进行乳腺生物反应器的研究奠定了分子生物学基础。     

军牧1号白猪、杜洛克猪和藏猪的氟烷基因和酸肉基因多态性分布

为了研究军牧1号白猪、杜洛克猪和藏猪氟烷基因和酸肉基因的多态性分布情况,试验采用PCR-RFLP方法对61头军牧1号白猪、51头杜洛克猪和51头藏猪的氟烷基因和酸肉基因多态性进行了检测。结果表明:军牧1号白猪的氟烷基因表现为单一的NN型;杜洛克猪的氟烷基因表现为多态性,等位基因N的频率为0.196 1,基因型分布符合哈代-温伯格平衡(χ2=3.033 9,P=0.081 5);藏猪的氟烷基因表现为单一的nn型。3个猪种的酸肉基因均表现为单一的rn/rn型。     

Q型烟粉虱芳香基硫酸酯酶B基因的克隆及表达分析

为明确烟粉虱Bemisia tabaci取食感染番茄黄化曲叶病毒(Tomato yellow leaf curl virus,TYLCV)的番茄植株后,其体内的芳香基硫酸酯酶B基因(arylsulfatase B,ARSB)是否能够做出应答反应,基于Q型烟粉虱基因组数据克隆得到ARSB基因cDNA全长,采用生物信息学方法分析其序列特征,并通过实时荧光定量PCR技术测定ARSB基因在Q型烟粉虱不同发育阶段、不同组织及携带TYLCV前后的表达量变化情况。结果显示:Q型烟粉虱ARSB基因的cDNA全长为1 731 bp,编码576个氨基酸,分子量为64.89 kD,具有ARSB的保守结构域。ARSB基因在Q型烟粉虱不同发育阶段均有表达,在卵期表达量最高,成虫期表达量最低;该基因在Q型烟粉虱头胸部的表达量显著高于腹部;Q型烟粉虱获取TYLCV 72 h后其体内ARSB基因表达量显著提高。表明ARSB基因在Q型烟粉虱不同龄期、不同组织内存在差异表达,并可能参与Q型烟粉虱对TYLCV的响应和传毒过程。     

Cloning of Proteinase Inhibitor Gene StPI in Diploid Potato and Its Expression Analysis

A full-length cDNA of proteinase inhibitor gene with completed open reading frame of 116 amino acids was cloned from Ralstonia solanacearum (Rs) resistant potato leaves using the rapid amplification of cDNA ends (RACE) method and designated as StPI. BLAST search against NCBI showed that the StPI gene shared 89% identity with potato proteinase inhibitor Ⅰ precursor in nucleotide and 74% in amino acid. Analysis of semi-quantitative RT-PCR indicated that this gene was induced by Rs as well as up-regulated by jasmonic acid (JA). The StPI gene expression reached the highest level during 6-12 h post Rs-inoculation or JA-treatment, and then leveled off. Moreover, this gene was strongly induced by JA and its mRNA accumulation increased more quickly than that of Rs-inoculation. The StPI gene may play a role in potato resistance against Rs. The induction of StPI by Rs invasion may have a similar signal transduction pathway with JA treatment.