正交设计优化青蒿SRAP-PCR反应体系及引物筛选

为了建立青蒿的SRAP最佳扩增体系,并筛选出SRAP多态性引物,本研究以青蒿叶片DNA为模板,采用正交试验设计,以Mg^2+、dNTP Mix、Taq DNA聚合酶、引物和DNA模板5种因素5个水平,对青蒿SRAP反应体系进行研究。结果表明,青蒿SRAP-PCR最佳反应体系为:引物0.6μmol/L、Mg^2+2.0 mmol/L、模板DNA 5.1 ng、Taq DNA聚合酶2.0 U、dNTPs 0.25 mmol/L,总体积为25μL。各因素对扩增反应均有不同影响,其中引物浓度的影响最大,dNTPs的影响最小。运用该体系对不同种质资源的青蒿进行验证,证明该体系稳定可靠,并在30个引物组合中筛选出了25对扩增条带清晰,多态性丰富的引物组合。这一结论为今后利用SRAP标记技术进行青蒿分子遗传学研究提供了科学依据。     

Introduction to the International Symposium on Lipid Science and Health and research progress in lipid science and health

The International Symposium on Lipid Science and Health (ISLSH), which was organized annually by Oil Crops Research Institute of Chinese Academy of Agricultural Sciences (OCRI-CAAS) since 2016, has gained a strong reputation and attracted hundreds of delegates from around the world for discussion of lipid research trends and advances every year, to promote research and academic exchanges in the fields of lipid science and health. The 5th International Symposium on lipid Science and Health was successfully held in Wuhan, China, from October 23rd to 25th, 2020, to celebrate the 60th anniversary of OCRI-CAAS. The two-day symposium gathered well-known experts specialized in lipid science to share the current state of lipid research with emphasis on aspects covering: (1) lipid profiling and characterization, (2) lipid preparation and modification, (3) lipid improvement and regulation, and (4) lipid nutrition and health. The symposium was conducted by a combination of on-site and network meeting. More than 250 distinguished delegates from academia and industry participated in the on-site multidisciplinary meeting, and thousands of scholars attended the virtual event. This paper is as a record of the symposium proceedings and a brief summary of the advances and trends in 4 aspects of lipid science and health.     

Cortical gene expression pattern in rat focal cerebral ischemia

AIM: To investigate the genes differential expression in cortex during rat focal cerebral ischemia.METHODS: cDNA microarray chips containing numerous cDNAs were used to investigate the gene expression pattern between samples of focal cerebral ischemia and sham-control operation rats. RESULTS: Two hundred and eleven genes differentially expressed were screened out, among these genes, up-and down-regulated genes were 199 and 12, respectively. CONCLUSIONS: The analysis of gene expression pattern of focal cerebral ischemia based on cDNA microarray can realize high-throughput screening of the genes associated with the focal cerebral ischemia. The differential expression of genes may be related to the pathogenesis of focal cerebral ischemic diseases.     

棉花蔗糖合成酶基因家族在纤维发育期时空表达模式分析

蔗糖合成酶(Sucrose Synthase)是调控植物糖类代谢的关键酶之一,在植物生长发育及渗透调节过程中起着重要的作用。该酶为多基因家族,各成员在植物不同组织及不同发育阶段发挥不同的作用。陆地棉(Gossypium hirsutum)中有15个Sus成员,本研究利用转录组高通量测序及实时定量PCR技术,检测了该基因家族的表达模式。结果表明,Gh Sus1D、Gh Sus3A、Gh Sus3D和Gh Sus6D在起始期(0~3 DPA)大量表达;GhSus7D在10 DPA的纤维中表达量快速增加,达到同期突变体胚珠的4.7倍;Gh Sus1A、Gh Sus5D在纤维伸长期(5~15 DPA)表达量显著高于胚珠,分别在10 DPA、15 DPA时达到峰值;Gh Sus3A、Gh Sus3D、和Gh Sus6D在纤维发育(15 DPA)时表达量重新上升,且在20 DPA时仍处于较高表达水平。Sus基因家族组织表达模式分析表明,它们在陆地棉品种徐州142野生型和突变体中的组织表达模式基本相同,根、茎、叶、花中没有显著差异。但不同Sus基因具有不同的组织表达特异性。     

Effect of water deficiency on relationships between metabolism,physiology,biomass,and yield of upland cotton(Gossypium hirsutum L.)

Drought is a common abiotic stress that considerably limits crop production. The objective of this study is to explore the influence of water deficiency on the yield, physiologic and metabolomic attributes in upland cotton cultivars(Gossypium hirsutum L). Cotton cultivars, 'Ishonch' and 'Tashkent-6' were selected to study the relationships among their physiologic, metabolomic and yield attributes during water deficiency. Deficit irrigation was designed by modifying the traditional watering protocol to reduce water use. Results indicate that cotton cultivars respond differently to water deficit stress. Water deficit significantly influenced plant height, the number of internodes, and sympodial branches in both cultivars. However, yield components such as the number of bolls, boll seed, lint mass, and individual plant yield were significantly reduced only in 'Tashkent-6'. The leaf area decreased and the specific leaf weight increased in 'Ishonch' under deficit irrigation conditions. However, 'Tashkent-6' demonstrated significant water loss compared to 'Ishonch', and both cultivars showed reduced transpiration rates. Untargeted metabolite profiles of leaves showed clear separation in 'Ishonch', but not in 'Tashkent-6' under deficit irrigation compared to full irrigation. The individual metabolites such as proline and galactinol showed strong association with yield under water deficit stress. Moreover, this study indicates that leaf area and transpiration intensity influence yield during water deficiency. In summary, the correlation among morpho-physiologic, metabolic, and yield components significantly varied between the two cultivars under water deficiency. The flowering stage was sensitive to water stress for both cultivars. The direct relationship between physiology, metabolism, and yield may be a useful selection criterion for determining candidate parents for cotton drought tolerance breeding.     

基于亲水相互作用液相色谱-三重四极杆质谱法研究白茶萎凋过程中代谢物的变化

采用亲水相互作用液相色谱-三重四极杆质谱法,对白茶鲜叶(F)和室内萎凋处理29 h(W)、53 h(P)的2个萎凋样进行了代谢物靶向检测,Quality control(QC)样本中共检测到111个代谢物,主要包括47种氨基酸及其衍生物、29种核酸类代谢物、12种维生素和辅酶、17种糖代谢物和6种甘油磷脂代谢物。与鲜叶F相比,萎凋叶W和P中显著增加的代谢物分别有39种和41种,其中共有代谢物35种;显著降低的代谢物分别有9种和11种,其中共有代谢物8种。显著变化代谢物分析结果表明,萎凋叶的细胞膜受损、DNA和RNA降解、氨基酸衍生物显著增加、糖代谢异常。根据代谢物含量的变化幅度和组成特异性,筛选了表征萎凋叶细胞膜生理状态和核酸、糖代谢的标志代谢物。此外,针对代谢物中氨基酸和核苷酸的组成特点,提出了研究氨基酸衍生物滋味特性的必要性和应用代谢谱分析建立萎凋工艺适度标准的可能性。     

四川省不同地域的核盘菌遗传多样性分析

 核盘菌(Sclerotinia sclerotiorum)属于世界性分布的植物病原真菌,可以危害油菜等多种经济作物。研究不同地域核盘菌的遗传多样性对了解核盘菌的遗传演化过程和指导病害防控具有重要意义。实验采用序列相关扩增多态性(sequence-related amplified polymorphism,SRAP)标记对四川省17个不同地理来源的66株核盘菌菌株的遗传多样性进行了分析。10对检测引物共获得129个位点,其中123个为多态位点,占95.35%。UPGMA聚类结果显示,在相似性系数为0.7时,66个核盘菌菌株分为5类(Ⅰ、Ⅱ、Ⅲ、Ⅳ、Ⅴ),分别包含60、2、2、1和1个菌株。在相似性系数为0.74时,第Ⅰ类又可分为3个亚类(Ⅰ-1、Ⅰ-2、Ⅰ-3),分别包含21、37和2个菌株。聚类及组成分分析结果显示,四川省各地区的核盘菌菌株具有较高的遗传多样性,但其遗传变异与菌株地理来源无明显相关性。     

温度胁迫下沙葱萤叶甲小热激蛋白基因GdHsp20.6的克隆及表达分析

为明确小热激蛋白在沙葱萤叶甲Galeruca daurica应对温度胁迫中的作用,采用PCR方法克隆沙葱萤叶甲小热激蛋白基因Hsp20（GdHsp20.6）完整的开放阅读框（open reading frame,ORF）序列,应用在线软件对GdHsp20.6基因进行生物信息学分析,通过原核表达技术诱导表达及纯化其编码蛋白,并通过实时荧光定量PCR（real-time quantitative PCR,RT-qPCR）技术分析不同温度胁迫下GdHsp20.6基因的表达量。结果显示,GdHsp20.6基因ORF序列长度为543 bp,编码180个氨基酸,预测分子量为20.6 kD,无跨膜区和信号肽。GdHsp20.6氨基酸序列有高度保守的α-结构域。GdHsp20.6氨基酸序列与其它鞘翅目昆虫的Hsp20氨基酸序列有较高的一致性,其中与花绒寄甲Dastarcus helophoroides的Hsp20.99氨基酸序列的一致性最高,为63%。GdHsp20.6基因在大肠杆菌Escherichia coli BL21（DE3）细胞系中成功表达,经异丙基-β-D-硫代吡喃半乳糖苷（isopropyl-β-d-thiogalactoside,IPTG）诱导后GdHsp20.6蛋白成功表达,十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（sodium dodecyl sulfate-polyacrylamide gel electrophoresis,SDS-PAGE）和Western-blot分析表明融合蛋白大小与预测大小一致,并纯化获得了纯度较高的目的蛋白GdHsp20.6。低温（-10~5℃）和高温（35~40℃）处理1 h以及处理后25℃恢复30 min均能诱导GdHsp20.6基因表达上调,并且0℃处理30~120 min也能诱导GdHsp20.6基因表达上调。表明GdHsp20.6基因在沙葱萤叶甲应对低温和高温胁迫中可能起着重要作用。