硫腺苷甲硫氨酸合成酶基因的克隆及其在盐胁迫条件下的不同表达

硫腺苷甲硫氨酸作为甲基供体在转甲基反应中起到重要作用.为了解硫腺苷甲硫氨酸在盐地碱蓬(Suaedasalsa (L.)Pall)耐盐中的作用,我们对可能编码硫腺苷甲硫氨酸合成酶的基因(SsSAMS2)进行了分析.该基因在经400 mmol/L NaCl处理的盐地碱蓬地上部分的λ-Zap cDNA文库中克隆到,其插入片段全长1 531 bp,包含一个395个氨基酸的开放阅读框架,该基因推断的分子量约为43 kD.SsSAMS2与长春花(Catharanthus roseus)的SAMS2在氨基酸水平上的一致性为93%.Southern杂交显示,SsSAMS2在盐地碱蓬基因组中可能是两个拷贝.Northern分析显示硫腺苷甲硫氨酸合成酶基因受NaCl等胁迫的正调控.酶活性检测表明,NaCl胁迫条件下该酶活性增强.     

盐胁迫下过量表达腺苷甲硫氨酸合成酶基因的转基因烟草的生长

以转腺苷甲硫氨酸合成酶基因（SsSAMS2）烟草纯合子ST8-9为实验材料,研究盐胁迫下过量表达SsSAMS2对转基因烟草生长影响的结果表明,200mmol·L-1NaCl处理后的转基因烟草的光合速率和生物量都比野生型烟草高,积累的自由多聚胺比较多,同时精氨酸脱羧酶的转录本也更丰富。显示转基因烟草的耐盐性比野生型烟草高,多聚胺在烟株缓解盐胁中可能是起了重要作用。     

过量表达腺苷甲硫氨酸合成酶基因能提高转基因烟草中多聚胺的生物合成

以土壤农杆菌介导的转化方法将盐地碱蓬的腺苷甲硫氨酸合成酶cDNA（SsSAMS2）转化到烟草K326中。PCR分析的结果表明,SsSAMS2整合进K326的基因组内。共筛选到8个转基因纯合品系,分析其中3个品系（ST8．9、ST14—2、ST3．5）基因表达和多聚胺含量的结果表明,SsSAMS2可在转基因烟草中表达,转基因烟草中的多聚胺含量明显高于野生型烟草。这些结果表明,腺苷甲硫氨酸合成酶基因已在转基因烟草中过量表达并导致多聚胺含量提高。     

盐地碱蓬(Suaeda salsa)APX 基因的克隆及盐胁迫下的表达

从盐地碱蓬 (Suaedasalsa)中克隆了抗坏血酸过氧化物酶 (ascorbateperoxidase ,APX)的全长cDNA(SsAPX) ,基因注册号为AY0 34 893。SsAPX全长 1.1kb ,推导的氨基酸序列长为 2 5 0个氨基酸残基。BLAST同源性分析表明 ,该cDNA与已报告的菠菜(Spinaciaoleracea)细胞质抗坏血酸过氧化物酶基因同源性最高 ,在核苷酸水平上一致性为 87% ,在氨基酸水平上一致性为 89%。Southern杂交表明APX基因在盐地碱蓬基因组中只有 1个拷贝。盐 (NaCl 40 0mmol/L)处理不同时间后的Northern杂交分析表明盐地碱蓬中SsAPX基因在盐胁迫下表达量增加 ,而且在盐胁迫下抗坏血酸过氧化物酶的活性也显著地增加 ,说明该基因受盐诱导。推测抗坏血酸过氧化物酶可能在保护盐地碱蓬免受氧化损伤的过程中起到一定作用     

红树植物杯萼海桑SAMS基因的克隆与生物信息学分析

红树植物杯萼海桑是最耐盐的红树植物之一。S-腺苷甲硫氨酸合成酶(S-adenosylmethionine synthetase,SAMS)是S-腺苷甲硫氨酸(S-adenosylmethionine,SAM)生物合成途径的关键酶。SAMS作为一个逆境胁迫响应蛋白在植物的耐盐调控中发挥着极其重要的作用。本文结合杯萼海桑根的转录组注释,根据编码区序列设计引物,通过PCR克隆杯萼海桑SAMS基因的编码区cDNA,并对其进行生物信息分析,为研究杯萼海桑适应逆境的机制奠定理论基础。结果显示PCR扩增了一个长1 182 bp的基因片段,该片段编码由393个氨基酸组成的S-腺苷甲硫氨酸合成酶。同源性比对及进化树分析显示杯萼海桑的SAMS氨基酸序列进化上相对保守。本研究首次从红树林植物杯萼海桑中克隆S-腺苷甲硫氨酸合成酶基因,并获得其编码区序列,为进一步研究杯萼海桑应对逆境胁迫的分子生物学机制与胁迫相关基因调控网络奠定基础。     

烟草中一条新的S-腺苷甲硫氨酸合成酶基因的克隆及表达分析

利用差异显示技术从烟草中分离到一个受乙烯利诱导表达的cDNA片段,结合RACE扩增技术获得该基因的全长序列。该全长cDNA共含1350个碱基,编码区有1170 bp,通过GenBank同源性比较发现它与一条已知的烟草S-腺苷甲硫氨酸合成酶(SAMS1,EC2．5．1．6)的基因编码区存在90％的同源性,编码的氨基酸顺序96％同源,而在cDNA的5’及3’非编码区同源性很低,且有一些短片段的缺失。设计合适的引物分别对这两个SAMS基因进行RT-PCR分析表明,SAMS1基因受高温诱导表达增强,但不受甲基紫晶诱导的氧化胁迫及盐胁迫影响,而新分离到的这个基因(SAMS2)的表达同时受高温、甲基紫晶及高盐3种胁迫的诱导。     

盐地碱蓬(Suaeda salsa)中SsINPS基因的分子克隆及表达特性分析

肌醇 1 磷酸 (I 1 P)合成酶 (EC5 .5 .1 .4,INPS)是肌醇生物合成中的关键酶 ,催化葡萄糖 6 磷酸 (G 6 P)到I 1 P的反应。从该实验室已构建的NaCl40 0mmol/L处理的盐地碱蓬 (Suaedasal sa)cDNA文库中克隆了肌醇 1 磷酸合成酶的全长cDNA (S .salsamyo inositol 1 phosphatesynthase,SsINPS) ,基因注册号为AF43 3 879。SsINPS全长约 1 986bp ,含有开放式阅读框架 1 5 3 0bp ,3′和 5′的非翻译区分别为 1 3 9bp和 3 1 7bp ;推导的氨基酸序列全长 5 1 0个氨基酸残基 ,分子量约为 5 6 .7kD ,pI值为 5 .3 5。BLAST同源性分析表明 ,该cDNA与已报告的冰叶日中花 (Mesembryanthemumcrys tallinum)的INPS基因同源性最高 ,其中 ,核苷酸水平的同源性为 91 % ,氨基酸水平上的同源性为84%。以SsINPS全长cDNA为探针进行的South ern杂交结果表明 ,SsINPS基因在盐地碱蓬基因组中只有一个拷贝 ;Northern结果表明 ,在盐处理(40 0mmol/L的NaCl)下 ,SsINPS在叶中的表达量有显著的增加。从而说明SsINPS在盐胁迫下是上升调节的     

盐地碱蓬过氧化氢酶基因的克隆及盐胁迫下的表达分析

过氧化氢酶是清除H2O2的重要酶类.从400 mmol/L NaCl处理的盐地碱蓬(Suaeda salsa(L.)Pall)地上部分的cDNA文库中克隆了两个编码过氧化氢酶的cDNA(Sscat1和Sscat2),其中Sscat1(1.7kb)是一个全长cDNA克隆,编码一个492个氨基酸的开放阅读框架,而Sscat2(1.1kb)是一个cDNA片段.据编码Sscat1 3＇端的287个氨基酸的cDNA序列与Sscat2的cDNA序列进行的BLAST同源性分析表明,Sscat1和Sscat2在核苷酸水平的一致性为71.9%,在氨基酸水平上的一致性为75%.Southem杂交表明,Sscat1在盐地碱蓬基因组中为多拷贝基因,Sscat2则为一个单拷贝基因.Northern杂交结果表明在盐胁迫条件下Sscat1和Sscat2的表达存在差异:400 mmol/L NaCl处理48h的盐地碱蓬根中的Sscat1和Sscat2 mRNA水平比对照显著提高,但是在叶中仅Sscatl受盐诱导表达.不同盐处理时间下的表达分析也证实,在盐地碱蓬叶中仅Sscat1受盐诱导表达.这说明Sscat1和Sscat2在盐地碱蓬中是差异调控的.生理分析表明过氧化氢酶的活性在盐胁迫条件下显著提高.     

腺苷甲硫氨酸合成酶的提取纯化研究

腺苷蛋氨酸具有转甲基、转硫和转氨丙基等重要生理作用,已成为治疗疾病的重要药物。目的：为腺苷蛋氨酸合酶的基因克隆做准备。方法：研究了腺苷甲硫氨酸合成酶的提取和纯化。腺苷蛋氨酸合酶为胞内酶,其提取需先进行细胞破碎,然后进行盐析和离子交换层析等方法来纯化。酵母的破壁试验考察了研磨、加入有机溶剂和超声波等不同的破碎方法。结果：超声波破碎法最好,得到粗酶液的酶活力为0．934U／ml;经过硫酸铵盐析后,利用离子交换层析法纯化腺苷甲硫氨酸合成酶,作出了腺苷甲硫氨酸合成酶的穿透曲线和洗脱曲线。     

盐地碱蓬叶片液泡膜H+-ATPase B亚基的克隆及盐胁迫下表达分析

植物液泡膜H -ATPase在建立跨液泡膜质子梯度、促进液泡Na 区域化、提高植物耐盐性方面发挥着重要作用.本实验从盐生植物盐地碱蓬(Suaeda salsa L.)cDNA文库分离到碱蓬叶片液泡膜H -ATPase B亚基cDNA克隆.测序表明该基因长达1 974 bp,开放阅读框有1 470 bp编码489个氨基酸,含有一个保守的ATP结合位点,其蛋白分子量约为54.29 kD.Northem及Western印迹表明盐地碱蓬液泡膜H -ATPase B亚基表达明显受NaCl胁迫诱导,并且在NaCl胁迫下,B亚基在转录及翻译水平上与液泡膜H -ATPase c亚基存在协同作用.盐胁迫下,盐地碱蓬液泡H -ATPase B亚基与c亚基的协同表达增加了液泡H -ATPase的数量,从而提高了液泡H -ATPase活性,为碱蓬叶片液泡Na 区域化提供了动力,最终提高了碱蓬植株的耐盐性.     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted.     

Comparative morphology of the carpel in the Liliaceae: Hewardieae, Petrosavieae, and Tricyrteae

The young pistils in the melanthioid tribes, Hewardieae, Petrosavieae and Tricyrteae, are uniformly tricarpellate and syncarpous. They lack raphide idioblasts. All are multiovulate, with bitegmic ovules. The Petrosavieae are marked by the presence of septal glands and incomplete syncarpy. Tepals and stamens adhere to the ovary in the Hewardieae and the Petrosavieae but not in the Tricyrteae. Two vascular bundles occur in the stamens of the Hewartlieae and Tricyrtis latifolia. Ventral bundles in the upper part of the ovary of the Hewardieae are continuous with compound septal bundles and placental bundles in the lower part. Putative ventral bundles occur in the alternate position in the Tricyrteae and putative placental bundles in the opposite. position in the Petrosavieae. The dichtomously branched stigma in each carpel of the Tricyrteae is supplied by a bifurcated dorsal bundle.     

肝癌中HBV和HCV基因和抗原的分布及意义

采用原位分子杂交方法检测HCV RNA及HBV X基因;采用免疫组织化学方法研究HCV核心抗原,非结构区C33c抗原及HBxAg在肝细胞肝癌中的定位及分布.结果表明(1)HCV RNA、HBV X基因在肝细胞肝癌组织检出率分别为40%(55/136)和82%(112/136).HCV RNA定位于癌细胞的胞浆内,阳性细胞呈散在、灶状及弥漫分布三种形式;HBV X基因在肝癌细胞中的分布呈胞浆型、核型及核浆型,阳性细胞也呈上述三种分布形式;(2)HCV C33c抗原、核心抗原在肝细胞肝癌中的阳性率为81%(133/164)及86%(141/164).C33c抗原定位于癌细胞及肝细胞的胞浆内;核心抗原既定位于癌细胞核中,又可定位于胞浆中.C33c抗原阳性细胞以灶状分布为主;而核心抗原阳性细