油菜黑胫病菌T-DNA插入诱变因素优化及突变体筛选

油菜黑胫病是由Leptosphaeria biglobosa引起的一种真菌病害。这种病害在我国油菜产区广泛发生，并造成一定的经济损失。为通过获取突变体来研究L. biglobosa的生态适应性机制及致病机制，本文优化了影响农杆菌介导转化（ATMT）油菜黑胫病菌L. biglobosa菌株Lb731的因素，评估转化子质量，并筛选相关突变体。结果明确了农杆菌介导转化菌株Lb731的最佳因素：潮霉素B浓度为50 μg/mL，转化受体（分生孢子）培养时间为15 d (20℃)，浓度为2 × 10^7~8孢子/mL，农杆菌-受体共培养温度为25℃，共培养时间为72 h。在最适条件下的转化效率达到80个转化子/百万分生孢子。T-DNA插入基因组的频率为100%，单拷贝插入频率为72.7%，转化子抗潮霉素性状能稳定遗传。从2136个转化子中获得了11个菌丝生长减缓突变体，7个色素产生缺陷突变体和14个分生孢子产生缺陷突变体，并从这些突变体中鉴定出7个致病力丧失突变体。采用hiTAIL-PCR技术，从3个突变体中获得了T-DNA插入位点侧翼序列。上述结果为深入研究L. biglobosa的生态适应性机制及致病机制提供了材料和线索。     

油料作物油脂合成调控研究进展

植物油脂不仅是人类可食用油的主要来源,也是人类生产生活中重要的可再生原料。本文概述了植物油脂的生物合成途径,从母体效应、QTL、GWAS等多个方面总结了油料作物油脂合成的遗传学研究进展,同时探讨了已知的油脂合成调控相关基因的功能。本文综述了该领域的研究现状,为深入了解油料作物油脂合成调控网络提供了参考,也为油料作物的分子改良和遗传育种提供了理论基础。     

SSCP Marker Development and Analysis of Candidate Restorer Gene Rf1 for Cotton Cytoplasmic Male Sterility

[Objective] Locating the cotton cytoplasmic male sterility (CMS) restorer gene Rf₁ is important for investigating restorer gene mechanisms and improving restorer lines. In our previous study, a gene cluster, with nine Pentatricopeptide repeat(PPR) genes and nine other genes, was found within the 160-kb Rf₁ target region in Scaffold 333. The objective here was to improve the density of Rf1-linked markers in the target region and determine the expression profiles of candidate genes. [Method] Using the sequences of the 18 genes, we designed 155 single-strand conformation polymorphism (SSCP) primers covering all of the gene sequences to identify the polymorphic SSCP markers between the fertile and sterile pools. Additionally, real-time polymerase chain reaction(PCR) was performed to analyze the expression profiles of eight candidate genes in the four developmental stages of buds of sterile, maintainer and restoring lines, respectively. [Result] In total, 15 polymorphic primers were identified. A genotype analysis of the F₂ population was conducted using the 15 primers and 3 other polymorphic simple sequence repeats (SSR) markers. The markers were distributed in a 4.8 cM range. In addition, owing to the influence of sterile cytoplasm or restorer genes, most of the genes showed different expression patterns in the four developmental stages of the three lines' buds. [Conclusion] SSCP markers tightly linked to Rf₁ were identified and the expression profiles of candidate genes were determined. This study provides a basis for the further fine mapping of restorer genes and for candidate gene screening.     

翠竹及菲白竹LEA3同源基因的克隆及序列分析

竹类植物在中国有着非常悠久的栽培历史,并被广泛应用于园林、造林、庭院以及食品等行业。本研究以翠竹和菲白竹为材料提取其叶片总DNA,采用PCR方法获得翠竹SpLEA3基因与菲白竹SfLEA3-1、SfLEA3-2基因;其中SpLEA3全长804 bp,编码195个氨基酸,GC含量为68.4%;SfLEA3-1全长803 bp,编码195个氨基酸,GC含量为68.2%;SfLEA3-2全长557 bp,编码144个氨基酸,GC含量为67.8%。通过ProtParamy等生物软件分析,SpLEA3、SfLEA3-1和SfLEA3-2与贵州悬竹LEA3基因同源性高达61%以上,且3个基因编码蛋白均属于亲水性蛋白。源自2个竹种LEA3基因均包含一个完整的开放阅读框,其编码的氨基酸含有4~6个由11个氨基酸组成的保守基元序列。本研究不仅为深入了解竹类植物抗旱的分子机理研究提供了基础数据,也为竹类植物的抗旱育种后续研究提供了科学依据。     

南瓜银叶突变体48a的表型特征及其遗传学分析

南瓜银叶突变体48a是在嫩食型中国南瓜中分离筛选到的稳定遗传自交系,银色叶不仅可以作为标记性状应用到育种中,还可为南瓜抗虫、抗病、耐寒等一系列研究提供重要的材料基础。笔者对银叶突变体的表型特征及叶片的解剖结构进行鉴定分析,发现植株整体长势、熟性与野生型无明显差异;成熟叶片正面全部呈银灰色,叶绿素含量明显降低,叶片上表皮细胞与栅栏细胞间明显剥离,存在明显的空隙。利用突变体48a和野生型49a南瓜自交系构建的六世代遗传群体,调查发现F2的绿叶与银叶符合3∶1的分离比,回交群体BC1P1分离比符合1∶1,表明南瓜银色叶性状由单隐性基因控制。     

Molecular identification of blast resistance genes in rice landraces from northeastern India

Rice blast disease caused by the fungus Magnaporthe oryzae is one of the most devastating diseases causing huge losses worldwide. In the present study, major blast resistance genes were investigated in landraces originating from northeastern India. Based on phenotypic evaluation, 288 landraces were classified into three distinct groups: resistant (75), moderately resistant (127) and susceptible (86). The genetic frequencies of the 18 major blast resistance genes were between 6.2% and 27.4%, with only two genotypes possessing a maximum of nine blast resistance genes. The cluster and population structure analysis grouped the landraces into two groups. Through principal coordinate analysis, the scatter plots partitioned the resistant and moderately resistant landraces into different groups. Analysis of molecular variance showed maximum (96%) diversity within populations and least (4%) diversity between populations. Association analysis identified six markers, CRG4_2, RM72, tk59-2, pi21_79-3, RM1233 and RM6648, that are significantly associated with blast disease and explained a phenotypic variance of 1.1–6.5%. The associated genes could be used in marker-assisted rice breeding programmes for gene pyramiding to develop rice varietal resistance against blast disease. The present study represents a valuable blast resistance genetic resource that could be used for identification of new R genes, donors for blast resistance, and genomic studies.     

Construction and Partial Biological Characteristics Analysis of Listeriolysin S llsB Gene Deletion Strain

In order to analyze the effect of listeriolysin S (LLS) llsB gene deletion on the biological characteristics of Listeria monocytogenes (LM),this study used homologous recombination to construct the llsB gene deletion strain LM90-ΔllsB,and the biological characteristics of growth characteristics,median lethal dose (LD₅₀) and organ-borne bacteria were studied in healthy Kunming male mice at 8 weeks old and weighing 40 g±5 g.The llsB gene deletion strain was successfully constructed,and the deletion strain had good genetic stability through continuous passage to 20 generations in vitro.Based on its growth curve examination,we found that the growth rate of the mutant strain was slightly higher than that of the parent strain.The results of mice infection test showed that the LD₅₀ of the parent strain and the deletion strain were 10^6.17 and 10^6.50 CFU,respectively.Compared with the parent strain,the amount of bacteria load of the deletion strain in the liver and spleen of the mice was extremely significantly decreased (P<0.01),the results showed that the infection ability of the mutant strain on mice was obviously weakened.No Listeria monocytogenes was detected in the brain.The results suggested that llsB gene might have direct or indirect regulatory effect on some biological characteristics of LM90,and it would provide a theoretical basis for further understanding of the pathogenic mechanism of LLS,prevention and control of listeriosis.     

茶树叶片响应茶饼病侵染的转录组分析

采用高通量测序技术对被茶饼病病菌侵染的茶树叶片进行转录组测序,筛选得到差异基因359个,其中248个上调表达,111个下调表达。差异基因中有216个获得GO（Gene ontology）数据库功能注释,主要涉及到生物合成过程、催化活性、细胞过程等诸多生理生化过程;KEGG（Kyoto encyclopedia of genes and genomes）数据库富集分析发现,共有106个基因被注释到47个代谢通路中,其中,单萜生物合成、卟啉和叶绿素代谢、核糖体、氮代谢、双萜生物合成、植物病原互作等通路显著富集。有32个差异基因被鉴定为转录因子,分布在16个转录因子家族中。利用实时荧光定量PCR（Real time quantitative PCR,qRT-PCR）验证了随机挑选的差异基因在感病叶片和未感病叶片中的相对表达量,与转录组测序结果的变化趋势一致。结果表明,茶树响应病原菌侵染是一个复杂的过程,大量基因被诱导或抑制表达,与抗病相关的转录因子被大量激活且上调表达。本研究为深入挖掘茶树抗病基因及进一步研究抗病分子机制提供了理论依据。