营养与株型调控对雾培马铃薯的结薯生理影响

本研究采用3套营养液配方与3种株型调控措施探讨了不同马铃薯(Solanum tuberosumL.)品种的结薯生理效应。结果表明:不同营养液配方对雾培马铃薯的“源流库”皆存在一定的影响;采用以前期最小、结薯期最大的N/K和NH4 -N/NO3--N比值的A2营养液,对植株的光合势、叶绿素含量、根系活力、匍匐茎数、总薯数、总薯重以及后期末茎叶鲜重与干重等的增长刺激作用最大,其中又以“CIP24”品种的结薯性能最好;下放植株、喷施矮壮素和剪尖处理的不同组合均能使“CIP24”品种在A2营养液下增加光合势,降低株高,增加鲜重和增大根系体积。     

用生物素标记核酸探针检测鸭瘟病毒DNA的研究

用生物素-11-脱氧尿苷三磷酸(Biotin-11-dUTP),通过缺口平移法标记提纯的鸭瘟病毒 DNA,制备生物素化鸭瘟病毒 DNA 全基因组探针;以斑点杂交检测固定在硝酸纤维膜上的样品鸭瘟病毒 DNA 同源序列,杂交后用亲和素-碱性磷酸酶孵育,底物显色,阳性反应呈蓝紫色斑点。试验结果表明,生物素标记核酸探针可检出10pg 提纯的鸭瘟病毒DNA,并检出稀释10~5倍和肝组织鸭瘟病毒 DNA;对鸡马立克氏病毒 DNA,鸡痘病毒DNA 和噬菌体 DNA 无杂交反应;对鸭瘟病毒弱毒 DNA 产生微弱杂交。该技术具有快速、敏感、特异和无放射污染等优点。     

TCS基因转化泡桐及抗病能力

泡桐(Paulownia)是经济价值较高的优良绿化造林树种.由于丛枝病的危害,每年我国林业的直接经济损失就高达数千万元,并且严重影响农民种植泡桐的积极性(杨俊秀等,2007).     

Promising genetic resources for resistance to wheat streak mosaic virus and the wheat curl mite in wheat-<Emphasis Type="Italic">Thinopyrum</Emphasis> partial amphiploids and their derivatives

Wheat streak mosaic virus (WSMV), vectored by the wheat curl mite (WCM),Aceria tosichella Keifer, is one of the most destructive viral diseases of wheat found in many wheat producing areas of the world. Host resistance is the most effective method for controlling this disease and its vector. Symptomatological analysis and enzyme-linked immunosorbent assay (ELISA) were used to characterize WSMV-resistance in wheat-alien partial amphiploid lines and their derivatives. The results showed that most of partial amphiploids derived fromThinopyrum ponticum andTh. intermedium were free of systemic symptoms with very low ELISA readings that were similar to that of the non-inoculated Chinese Spring control. While the partial amphiploid lines 693 and PWM706 were identified as new genetic resources of resistance to WSMV. The present study demonstrated that both symptomatological and ELISA methods efficiently assessed WSMV-resistance in the wheat-alien hybrids and systemic symptom incidence and ELISA absorbance readings were highly correlated (r ₂ = 0.8658–0.9323) over time following inoculation. The ELISA results also indicated that the virus did not buildup in the plant tissues of these virus-resistant partial amphiploids. Similar results were observed in chromosome translocation and substitution lines that have the geneWsm1 conferring WSMV resistance. However, the lines containing the geneWsm1 and all the partial amphiploid lines, except Agrotana, were susceptible to the WCM. One line derived from a cross of wheat and Agrotana, was effective in controlling the spread of WSMV and was highly resistant to the WCM. Another line and an accession ofTriticum dicoccoides (Koern.) Schweinf. were highly susceptible to WSMV and WCM. Early disease development was delayed in a new hard red winter cultivar McClintock. The partial WSMV-resistance of McClintock was demonstrated by initially low ELISA readings, and a lower percentage of infected plants than other WSMV-susceptible wheat. The use of the identified promising sources of resistance to WSMV and the WCM in wheat breeding is discussed.LRC Contribution No. 387-01061.     

应用RNA沉默技术获取抗黄瓜花叶病毒(CMV)和烟草花叶病毒(TMV)转基因烟草

RNA沉默是迄今最为有效的抗病毒策略,利用该策略不但能获得免疫转基因植株,且所得植株不易与其他病毒基因重组或异源包壳、生物安全性较高。将本实验室已构建的携带有烟草花叶病毒(TMV)部分移动蛋白基因(ΔMP)和黄瓜花叶病毒(CMV)部分复制酶基因(ΔRep)反向重复结构的植物表达载体pBIN438-MP-Rep(i/r),用农杆菌浸润法转化普通烟草品种K326,共得196株转化植株,经卡那霉素筛选和PCR检测发现128株为阳性转基因株;PCR-Southern和RT-PCR分析表明外源基因已整合到烟草基因组并在转录水平上得到表达;ELISA结果显示20.3%的转基因植株对CMV和TMV复合侵染表现免疫性。本结果为利用RNA沉默技术进行植物抗多种病毒育种提供重要数据,为防治其他多种病毒复合侵染提供借鉴。     

利用BSMV-VIGS技术快速分析小麦TNBL1基因的抗黄矮病功能

小麦黄矮病是由蚜虫介导的大麦黄矮病毒(Barley yellow dwarf virus, BYDV)侵染引起的小麦重要病害之一。利用cDNA-AFLP分析,筛选出在抗黄矮病小麦易位系YW642中特异表达的长度为292 bp的 cDNA片段,以此片段为启始序列,利用RACE和RT-PCR技术克隆出该基因的全长cDNA序列,推导该基因编码1个NBS-LRR蛋白,将其命名为TNBL1。本研究利用大麦条纹花叶病毒(BSMV)诱导的基因沉默(virus-inducing gene silencing, VIGS)技术,快速分析TNBL1是否参与小麦抗黄矮病反应。通过PCR添加酶切位点、定向酶切与连接,将TNBL1特异的292bp片段反向整合到BSMV-γ链的多克隆位点上,获得重组载体BSMV-γ: TNBL1as,体外转录BSMV-VIGS载体的3个组分(BSMV-TNBL1as、BSMV-α和BSMV-β),等量混合、摩擦接种到抗黄矮病的小麦易位系YW642幼苗叶片上,使YW642中TNBL1基因沉默,然后接种BYDV病原进行黄矮病抗性鉴定。结果表明,TNBL1基因沉默后的YW642对BYDV敏感、显现感病症状,其体内BYDV含量较未发生基因沉默的YW642中的明显增加,证明TNBL1基因是正向调控小麦抗BYDV反应的1个重要基因。     

抑制烟草花叶病毒侵染的植物提取物的筛选

用半叶枯斑法,在心叶烟上测试了33种植物乙醇提取物水溶液对烟草花叶病毒(Tobacco mosaicvirus,TMV)侵染的抑制效果。结果表明,雪松、桉树提取物能较好地抑制病毒(TMV)侵染,抑制率达到89.7%,78.3%;此外,菟丝子、红花蔷薇、桑、红千层、杜鹃、桂花、香石竹、滇厚朴、荠菜、芦荟等植物提取物也具有抑制烟草花叶病毒(TMV)侵染的效果,抑制率均在50%以上。     

长距离PCR法扩增RHDV全基因组及其序列分析

从人工感染RHDV致死的兔肝组织中提取病毒总RNA,应用特异性引物将病毒RNA反转录成cD-NA,以此为模板用长距离PCR法扩增RHDV全基因组,将成功扩增出的RHDV全基因组克隆到pMD18-T载体中,经PCR鉴定及酶切分析后进行序列测定。测序结果表明,JX97全基因组由7 437个核苷酸组成,包括由9个核苷酸组成5′NCR、59个核苷酸组成3′NCR和一个至少含有27个核苷酸的poly(A)尾。序列比较结果显示,RHDVJX97株的衣壳蛋白与已报道的参考毒株具有较高的序列同源性,均在90%以上,提示所有的RHDV分离株具有较近的亲缘关系。     

辣椒染病毒株与健株花药中游离脯氨酸含量和花粉萌发力的比较

对辣椒感染病毒病植株与健康植株花药中的游离脯氨酸和花粉萌发力进行了比较性研究,结果表明,从开花前7天到开花当天,病株花药中的游离脯氨酸含量均比同期健株的低,但以开花前4天到开花当天的差异较明显,开花前1天病株花药中游离脯氨酸含量比健株的低26.24％～39.96％,开花当天病株上的花粉萌发率比健株的低14.35％～33.31％。本研究结果从一个侧面反映了辣椒感染病毒病后在生理生化上发生的病理变化。