Satellite RNA reduces expression of the 2b suppressor protein resulting in the attenuation of symptoms caused by Cucumber mosaic virus infection

Satellite RNAs (satRNAs) depend on cognate helper viruses for replication, encapsidation, movement and transmission. Many satRNAs with different symptom modulation effects have been reported. The pathogenicity of satRNAs is thought to be the result of a direct interaction among the satRNA, helper viruses and host factors by unknown mechanisms. To understand the effect of satRNA of Cucumber mosaic virus (a severe field ShanDong strain, SD-CMV) on pathogenicity, and the possible involvement of host RNA silencing pathways in pathogenicity, we constructed biologically active CMV cDNA clones and a CMV-Δ2b mutant lacking the open reading frame of 2b, a silencing suppressor protein, in order to infect Nicotiana benthamiana and Arabidopsis with or without SD-satRNA. We found that SD-satRNA reduced the accumulation of the 2b protein and its coding RNA4A and attenuated the yellowing caused by SD-CMV infection. Small RNA analysis indicated that the 2b protein interfered with RNA silencing, specifically in the synthesis of CMV RNA3-derived small interfering RNAs (R3-siRNAs). The accumulation of R3-siRNAs in CMV-Δ2b infection was reduced in the presence of satRNA, for which greater accumulation of satRNA-derived siRNAs (satsiRNAs) was detected. Our results suggest that abundant SD-satRNA serving as target for RNA silencing may play a role in protecting helper CMV RNA, especially, subgenomic RNA4, from being targeted by RNA silencing. This compensates for the increase in RNA silencing resulting from the reduction in expression of the 2b suppressor in the presence of satRNA. Our data provide evidence that a plant silencing mechanism is involved in the pathogenicity of satRNA.     

The expression level of <Emphasis Type="Italic">Rosa Terminal Flower 1</Emphasis> (<Emphasis Type="Italic">RTFL1</Emphasis>) is related with recurrent flowering in roses

We examined the relationship between the recurrent flowering character and the expression patterns of TERMINAL FLOWER 1 (TFL1) homologs in roses, using flower buds of Rosa multiflora, R. rugosa, R. chinensis, and six other rose species and nine rose cultivars. RTFL1 (Rosa TFL1) genes were amplified from rose genomic DNA using a combination of degenerate and gene-specific primers by thermal asymmetric interlaced-PCR and normal PCR, respectively. Their copy numbers in different species were determined by Southern blots. We used real-time PCR to analyze the expression patterns of RTFL1 genes at four developmental stages (pre-sprouting, young, mid-aged, and mature flower buds). Our results show that there are at least three RTFL1 homologs in roses; RTFL1a, RTFL1b, and RTFL1c. The sequences of the homologs were more similar among the same homolog in different species than among the different homologs in the same species. For RTFL1a, we detected two copies in R. multiflora, two copies in R. rugosa, and one copy in R. chinensis. For RTFL1c, we detected one copy in R. multiflora, two copies in R. rugosa, and three copies in R. chinensis. We detected only one copy of RTFL1b in R. chinensis. RTFL1c was expressed at high levels at all four flowering stages in R. multiflora and R. rugosa, which are non-recurrent flowering species, whereas it was barely detected in R. chinensis (a recurrent flowering species) at any stage. These results were further verified in six other non-recurrent flowering species and nine recurrent flowering cultivars. These results suggest that the recurrent flowering habit in roses results from lower expression of RTFL1c, which may be related to recurrent flowering character in roses.     

Artificial MicroRNAs highly accessible to targets confer efficient virus resistance in plants

Short-hairpin RNAs based on microRNA (miRNA) precursors to express the artificial miRNAs (amiRNAs) can specifically induce gene silencing and confer virus resistance in plants. The efficacy of RNA silencing depends not only on the nature of amiRNAs but also on the local structures of the target mRNAs. However, the lack of tools to accurately and reliably predict secondary structures within long RNAs makes it very hard to predict the secondary structures of a viral genome RNA in the natural infection conditions in vivo. In this study, we used an experimental approach to dissect how the endogenous silencing machinery acts on the 3′ untranslated region (UTR) of the Cucumber mosaic virus (CMV) genome. Transiently expressed 3′UTR RNAs were degraded by site-specific cleavage. By comparing the natural cleavage hotspots within the 3′UTR of the CMV-infected wild-type Arabidopsis to those of the triple dcl2/3/4 mutant, we acquired true small RNA programmed RNA-induced silencing complex (siRISC)-mediated cleavage sites to design valid amiRNAs. We showed that the tRNA-like structure within the 3′UTR impeded target site access and restricted amiRNA-RISC-mediated cleavage of the target viral RNA. Moreover, target recognition in the less-structured area also influenced siRISC catalysis, thereby conferring different degrees of resistance to CMV infection. Transgenic plants expressing the designed amiRNAs that target the putative RISC accessible target sites conferred high resistance to the CMV challenge from both CMV subgroup strains. Our work suggests that the experimental approach is credible for studying the course of RISC target recognition to engineer effective gene silencing and virus resistance in plants by amiRNAs.     

羽叶南洋参的组织培养

1植物名称羽叶南洋参(Polyscias fruticosa var.plumata Bailey),别名南洋森和福禄桐。2材料类别茎段。3培养条件基本培养基为MS。芽诱导培养基:(1)MS 6-BA2mg·L-1(单位下同) NAA0.5。芽增殖     

农杆菌接种法作为一种简便的植物病毒载体的侵染方法

农杆菌接种法作为一种简便的植物病毒载体的侵染方法 贾洪革 庞永奇 方荣祥     

选择标记可去除的植物高效表达载体的构建

在常用的植物组成型表达载体pBI121的选择标记基因NPTII两侧插入同向的lox位点并用多克隆位点(MCS)取代了GUS基因序列,构建了NPTII基因可被去除的和可插入目的基因的通用植物表达载体pBI121-lox-MCS。替换pBI121-lox-MCS中驱动目的基因表达的35S启动子,可构建成一系列具有其他表达特性的植物表达载体,如本文描述的韧皮部特异表达载体pBdENP-lox-MCS。为方便地筛选去除选择标记基因的转基因植物,还构建了绿色荧光蛋白(GFP)表达框与NPTII表达框连锁的pBI121-gfp-lox-MCS载体。上述植物表达载体可广泛应用于培育选择标记可去除的转基因植物。     

了哥王的组织培养与植株再生

1植物名称了哥王[Wikstroemia indica（L.）C.A.Mey.],别名土木香、黄皮子、南岭荛花。2材料类别茎段。3培养条件基本培养基为MS。芽诱导培养基：     

针对leader序列的siRNA能够有效抑制SARS冠状病毒多个mRNA的表达

小干扰RNAs(siRNAs)能够有效降解具有互补序列的RNA.在SARS-CoV的基因组RNA和所有亚基因组RNA的5′端均有一段共同的leader序列,而且该leader序列在不同的病毒分离物中高度保守,因此leader序列可作为一个用于抑制SARS-CoV复制的有效靶点.研究表明,针对leader序列化学合成的siRNA和DNA载体表达的shRNA都可以有效抑制SARS-CoV mRNA的表达.Leader序列特异的siRNA或shRNA不仅可以有效抑制leader与报告基因EGFP融合基因的表达,而且还可以有效抑制leader与刺突蛋白(spikeprotein)、膜蛋白(membrane protein)和核衣壳蛋白(nucleocapsid protein)基因的融合转录产物的表达.结果表明,针对leader序列的RNA干扰可以发展成为一种抗SARS-CoV治疗的有效策略.     

3种除草剂对七星瓢虫的急性毒性与初级风险评估

为了评估除草剂对七星瓢虫Coccinella septempunctata的毒性影响和初级风险,采用药膜法进行了3种除草剂的急性接触毒性测试,并评估其初级风险。结果表明：10%噁唑酰草胺乳油、10%噁嗪草酮悬浮剂和20%异噁唑草酮悬浮剂对七星瓢虫的急性接触毒性分别为LR₅₀>3.61×10² g a.i./ha, LR₅₀>1.485×10² g a.i./ha, LR₅₀>3.60×10² g a.i./ha。10%噁唑酰草胺乳油、10%噁嗪草酮悬浮剂和20%异噁唑草酮悬浮剂的相对安全系数<5,属于中等及以下风险农药,对七星瓢虫的农田内和农田外喷雾场景风险均可接受(危害商分别为HQ_in<0.997,HQ_in<2.000,HQ_in<1.000,HQ_in均<5;HQ_off<0.020,HQ