粘虫核型多角体病毒一个新基因的序列和结构研究

本文报道ＴｓＮＰＶＤＮＡＥｃｏＲＶ／ＸｈｏＩ的５．５ｋｂ片段的克隆及其物理图谱。测定了其中一个８１９ｂＰ片段的全序列。在长３４６ｂｐ的完整ＯＲＦ的５’端除有ＴＡＴＡｂｏｘ和ＣＡＡＴｂｏｘ以外,还发现有典型的早期基因启动子元件ＡＣＧＴ和ＧＣ单元以及晚期基因的转录起始保守序列ＴＴＡＡＧ。预测的ＯＲＦ产物为１１４个氨基酸、分子量为１３ｋＤ的蛋白质,故命名为ｐ１３蛋白。在ｐ１３基因的Ｃ端和Ｎ端分别有一个亮氨酸拉链和两个类亮氨酸拉链结构（我们称为亮氨酸转型结构和ＬＶＴ重复结构〕．从终止密码起有一个双发卡环结构,ｐ１３基因的５’端调控单元及其排列与ＢｍＮＰＶ和ＡｃＮＰＶ的细胞序死抑制基因（ｐ３５）十分相似,但ｐ１３基因与已经发现的杆状病毒基因序列和氨基酸序列没有同源性．因此,这是一个新的早晚期基因,其调控元件可能具有重要功能。     

水稻矮缩病毒最外层外壳蛋白基因(S_2)cDNA克隆、序列分析及其在大肠杆菌中的表达

从水稻矮缩病毒（Ｒｉｃｅｄｗａｒｆｖｉｒｕｓ,ＲＤＶ）中国福建分离物中克隆分离了最外层外壳蛋白基因（Ｓ２）全长ｃＤＮＡ,并对其进行了序列分析,结果表明ＲＤＶＳ２ｃＤＮＡ全长３５１２ｂｐ,仅含一个３３４８ｂｐ的阅读框架,编码一个含有１１１６个氨基酸的蛋白（Ｐ２）。与基因库中已知基因序列比较,发现它与日本ＲＤＶＨ株系相应片段的核苷酸和氨基酸同源率分别为９４６％和９５４％,与轮状病毒ＶＰ２氨基酸序列有一定的同源性。Ｓ２核苷酸序列二级结构预测结果表明,５’端５０个核苷酸的二级结构为一个发夹结构和一个茎环结构。Ｐ２有４个富含亮氨酸的区域,位于Ｎ端亲水区域的１０个氨基酸（ＡＡ６９～７８）残基形成一个α螺旋,这些特点均与轮状病毒ＶＰ２的结构特征相似。ＳＤＳＰＡＧＥ和Ｗｅｓｔｅｒｎ印迹分析表明在大肠杆菌中分段高效表达了Ｓ２编码蛋白的Ｎ端和Ｃ端。     

黄瓜花叶病毒山东株(CMV-SD)RNA2全长cDNA克隆的构建及全序列分析

分别利用５’ＲＡＣＥ和３’ＲＡＣＥ确定了ＣＭＶＳＤＲＮＡ２的５’和３’末端序列,在此基础上,利用ＲＴＰＣＲ得到了ＲＮＡ２的５’端一半的ｃＤＮＡ克隆ｐＣ２５和３’端一半的ｃＤＮＡ克隆ｐＣ２３,并通过拼接构建了ＲＮＡ２全长ｃＤＮＡ克隆ｐＣ２Ｆ。通过对ｐＣ２５和ｐＣ２３进行序列测定,得到了ＲＮＡ２的全序列。序列分析结果表明ＣＭＶＳＤＲＮＡ２由３０４８ｎｔ组成,其中存在２个部分重叠的阅读框ＯＲＦ１（７９～２６５２ｎｔ）和ＯＲＦ２（２４１４～２７４６ｎｔ）,分别编码８５８ａａ的２ａ蛋白和１１１ａａ的２ｂ蛋白,并在２ａ蛋白的序列中发现了动植物病毒复制酶所特有的两个保守序列。该株系ＲＮＡ２核苷酸序列与分属ＣＭＶＩ亚组的Ｆｎｙ株系和ＩＩ亚组的Ｑ株系ＲＮＡ２的核苷酸序列同源性分别为９１７％和７５６％;２ａ蛋白的氨基酸序列同源性分别为９３８％和６７７％,２ｂ蛋白的氨基酸序列同源性分别为８３０％和５１３％。同源性比较的结果表明ＳＤ株系属于ＣＭＶＩ亚组。     

黄瓜花叶病毒山东株（CMV—SD）RNA2全和cDNA克隆的构建及全序列分析

分别利用５’ＲＡＣＥ和３’ＲＡＣＥ确定了ＣＭＶ－ＳＤＲＮＡ２的５’和３’未端序列,在此基础上,利用ＲＴ－ＰＣＲ得到了ＲＮＡ２的５’端一半的ｃＤＮＡＤＱ ＢＴＧ Ｐｃ２５和３’端一半的ｃＤＮＡ克隆ＰＣ２３,并通过拼接构建了ＲＮＡ２全长ｃＤＮＡ克隆ＰＣ２Ｆ。     

利用脊髓灰质炎病毒5‘NCR和RNA聚合酶基因可提高外源基因表达

将含脊髓灰质炎病毒（ＰＶ）ＲＮＡ聚合酶的不同长度基因片段克隆到载体ｐＳＧ５质粒上,分别构建了４个表达ＲＮＡ聚合酶的质粒。体外转录实验证明,ｐＳＧ５－ＰＯＬ１．９９和ｐＳＧ５－ＰＯＬ２．０３质粒转染细胞的提取物促进了特异的ＲＮＡ转录,表明两质闰可表达ＲＮＡ聚合酶。将ＰＶ的５’ＮＣＲ序列插在载体ｐＧＲＥＥＮ ＬＡＮＴＥＲＮ－１的ＣＭＶ启动子下游,构建了ｐＧＲＥＥＮ ＬＡＮＴＥＲＮ－１－５’ＮＣＲ质粒;     

黄鳝基因组中存在酪氨酸蛋白激酶受体基因同源序列

近年的研究表明。酪氨酸蛋白激酶受体通过其细胞膜外部的结构域与细胞外的信号分子配体结合后,激活本身位于细胞质内的激酶结构域。磷酸化的酪氨酸进而激活下游一系列信号分子。这些分子的激活引起细胞内基因表达的改变、最终导致细胞本身表型状态的变化。本文发现并研究了存在于鱼类中的酪氨酸蛋白激酶受体基因的同源序列ｍａｔｋ。实验用黄鳝和胡子鲇为集市采购。ＰＣＲ扩增采用人ＳＲＹ基因编码区的一对引物。分别为：５’ＣＣＣＧＡＡＴＴＣＧＡＣＡＡＴＧＣＡＡＴＣＡＴＡＴＧＣＴＴＣＴＧＣ３’和５’ＣＴＧＴＡＧＣＧＧＴＣＣＣＧＴＴＧＣＴＧＣＧＧＴＧ３’。分别制备雌雄性黄鳝基因组ＤＮＡ。进行ＰＣＲ扩增。２％琼脂糖凝胶电泳分析表明,雌雄性样品中均可见到约２５０ｂｐ的扩增带。将雄性的扩增产物ｍａｔｋ重组到ｐＵＣ１３载体上。对其进行测序。结果表明：ｍａｔｋ与人ＳＲＹ和ＳＲＹ盒基因序列无同源性,而与最近才报道的大鼠酪氨酸蛋白激酶受体基因ｐｔｋ３ｃＤＮＡ５’端序列具有５６％的序列一致性（图１）。有报导,人与鼠的酪氨酸蛋白激酶受体基因ＤＤＲ和ｐｔｋ３存在９６％的同源性。表明这种酪氨酸蛋白受体基因具有很强的保守性。以ｍａｔｋ为探针,对经过ＥｃｏＲＩ酶切过的     

转译优化对EPO基因在COS7细胞中表达的影响

利用ＣＯＳ７细胞暂时表达系统,研究转译起始序列对ＥＰＯ－ｃＤＮＡ表达的影响。通过ＤＮＡ重组技术,构建了原ＥＰＯ－ｃＤＮＡ表达载体ｐＣＳＶ－ＥＰＯ（１）,其转译起始序列为５＇ＡＡＴＴＣＡＴＧＧ３＇。同时通过定点突变技术,将起始序列改变成５＇ＣＣＡＣＣＡＴＧＧ３＇,而构建了另一表达载体ＰＣＳＶ－ＥＰＯ（２）。后经序列分析证明无误后和前均通过ＤＥＡＥ－ｄｅｘｔｒａｎ法转染ＣＯＳ７细胞上清,测定结果为     

兔2‘,5’—寡聚腺苷酸合成酶的诱导形成

通过对兔血细胞内２‘,５’－寡聚腺苷酸合成酶诱导形成的研究,发现聚肌苷酸与聚胞苷酸双链复合物只能诱导单核型细胞产生２‘,５－ＯＡＳＥ,而新城疫病毒（ＮＤＶ）、红细胞生成素（ＥＰＯ）可同时刺激ＰＢＭＣ和总红细胞中２’,５‘－ＯＡＳＥ的上升。实验证明总红细胞中２’－５‘－ＯＡＳＥ定位于网织细胞中,苯肼、ＮＤＶ和ＥＰＯ引起总红细胞中２’,５‘－ＯＡＳＥ活性的增加与外周血中网织细胞的增加有直接关系。注射ｐ     

Epstein—Barr病毒膜抗原在中国仓鼠卵巢（CHO）细胞中的表达

将切去３＇端穿膜序列的ＥＢ病毒膜抗原（ＭＡ）基因,插入ｐＳＶ２－ｄｈｆｒ质粒的ＳＶ４０早期启动子下游,构建了真核表达载体ｐＳＶ２－ｄｈｆｒＧＰＴＲ,使两个ＳＶ４０早期启动子分别调控ＭＡ和二氢叶酸还原酶（ｄｈｆｒ）基因。将该重组质粒转化ＣＨＯ－ｄｈｆｒ细胞。在选择培养基中筛选阳性克隆,用氨甲喋呤加压扩增,建立了表达ＥＢＶ－ＭＡ的克隆细胞系。Ｗｅｓｔｅｒｎ ｂｏｌｔ分析证明,所表达的蛋白的分子量大约为     

反义寡核苷酸对丙型肝炎病毒调控基因表达的体外抑制活性研究

为探讨反义寡核苷酸对丙型肝炎病毒的抑制活性,研究和开发新型抗ＨＣＶ药物。采用ＨＣＶ５’ＮＣＲ调控荧光素酶基因的稳定表达细胞株ＨｅｐＧ２．９７０６,评价了３条针对ＨＣＶ调控基因的ＡＳＯＤＮ,即ＨＣＶ３６３,ＨＣＶ３４９及ＨＣＶ２７９。将Ｌｉｐｏｆｅｃｔｉｎ包封的ＡＳＯＤＮ与ＨｅｐＧ２．９７０６细胞株每天作用５小时,连续３天后检测荧光素酶活性。     

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.     

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.     

Enterovirus 71 3C proteolytically processes the histone H3 N-terminal tail during infection

Highlights
1. The N-terminal tail of histone H3 is specifically cleaved during EV71 infection.
2. Viral protease 3C is identified as a protease responsible for proteolytically processing the N-terminal H3 tail.
3. Our finding reveals a new epigenetic regulatory mechanism for Enterovirus 71 in virus-host interactions.     

Detection of EBV and HHV6 in the Brain Tissue of Patients with Rasmussen’s Encephalitis

Rasmussen’s encephalitis (RE) is a rare pediatric neurological disorder, and the exact etiology is not clear. Viral infection may be involved in the pathogenesis of RE, but conflicting results have reported. In this study, we evaluated the expression of both Epstein-Barr virus (EBV) and human herpes virus (HHV) 6 antigens in brain sections from 30 patients with RE and 16 control individuals by immunohistochemistry. In the RE group, EBV and HHV6 antigens were detected in 56.7% (17/30) and 50% (15/30) of individuals, respectively. In contrast, no detectable EBV and HHV6 antigen expression was found in brain tissues of the control group. The co-expression of EBV and HHV6 was detected in 20.0% (6/30) of individuals. In particular, a 4-year-old boy had a typical clinical course, including a medical history of viral encephalitis, intractable epilepsy, and hemispheric atrophy. The co-expression of EBV and HHV6 was detected in neurons and astrocytes in the brain tissue, accompanied by a high frequency of CD8⁺ T cells. Our results suggest that EBV and HHV6 infection and the activation of CD8⁺ T cells are involved in the pathogenesis of RE.     

Perinatal Vertical Transmission of Chikungunya Virus in Ruili,a Town on the Border between China and Myanmar

正Dear Editor,Chikungunya virus (CHIKV), an arbovirus in the family of Togaviridae, genus Alphavirus, is transmitted by the A.aegyptii or A. albopictus mosquito, and causes disease in humans characterized by fever, rash, and arthralgia (Silva and Dermody 2017; Suhrbier 2019). It was first reported in 1953 in Tanzania, and caused only a few outbreaks and sporadic cases in Africa and Asia in last century. However, in the epidemic in 2004, CHIKV acquired mutations that conferred enhanced transmission by the A. albopictus mosquito(Schuffenecker et al. 2006). Since then, it has successively caused outbreaks in Africa, the Indian Ocean, South East Asia, the South America, and Europe (Zeller et al. 2016).     

Development of a Novel Reverse Transcription Loop-Mediated Isothermal Amplification Method for Rapid Detection of SARS-CoV-2

In conclusion, the novel visual RT-LAMP assay is a simple, rapid, and sensitive approach for detection of SARS-CoV-2, and it is ready for application in primary care and community hospitals or health care centers, and even patients' own houses in response to the current SARS-CoV-2 epidemic because the assay does not require sophisticated equipment and skilled personnel. Furthermore, it is also ready to be used in fields for screening samples from wild animals and environments to facilitate the identification of potential intermediate hosts that mediate the cross-species transmission of SARS-CoV-2 from bats to humans.