天麻AFLP分析技术体系的建立

目的:建立一个适于天麻研究用的AFLP分析技术体系。方法:以11份天麻种质资源为试材,构建供试天麻的AFLP指纹图谱,同时对AFLP分析过程中DNA提取、酶切、连接、预扩增、引物筛选、选择性扩增、电泳和银染等多种因素进行分析。结果:AFLP分析体系要求DNA模板质量高,酶切连接可同时进行,37℃、9h效果较好,预扩增要求高质量DNA聚合酶,产物15倍稀释作为选择性扩增模板效果好,制胶前玻璃板的处理和银染时间的掌控对获得较好结果非常关键。P-GGA/M-GT引物构建的指纹图谱拥有可清晰辨认的扩增条带45条。结论:AFLP指纹技术具有稳定性高、重复性好等优点,可用于天麻DNA分析。     

短蛸AFLP分子标记分析体系的优化与建立

本研究构建了短蛸扩增片段长度多态性(AFLP)分析体系,对DNA提取、双酶切反应、连接反应、预扩增反应、选择性扩增反应和银染等步骤进行了分析。得到了一种适于短蛸AFLP技术分析的优化体系,该体系中各优化因素为:模板DNA浓度为200 ng/μL;酶切体系中,MseI和EcoR I各加入5 units,缓冲液使用MseI buffer Tango,反应时间为3-4 h;连接最适反应时间为12 h;预扩增产物最适稀释倍数为20倍。该体系的构建为AFLP技术在短蛸分子遗传多样性研究中的应用奠定了基础。     

蚕豆AFLP技术体系的建立与优化

对蚕豆DNA提取质量和浓度、DNA双酶切与连接、酶切连接产物的预扩增和选择性扩增等AFLP技术体系中的关键技术进行了优化处理,构建了蚕豆AFLP银染技术体系。酶切与连接可在12.5μl体系中一步完成,酶切连接温度为37℃,反应时间12~14 h;预扩增体系为20μl,选择性扩增体系为10μl。采用该技术体系应用8对引物构建的蚕豆种质资源AFLP指纹图谱,扩增条带多、多态性强且质量好,可满足遗传多样性分析要求。     

华中五昧子AFLP反应体系的建立

目的:建立一个适于华中五味子研究用的AFLP反应体系.方法:以华中五味子硅胶干燥嫩叶为试材,采用改良CTAB法提取到高质量DNA.通过琼脂糖凝胶电泳和聚丙烯酰胺凝胶电泳对Mse I/EcoR Ⅰ双酶切、连接、预扩增和选择性扩增过程中的关键因素进行分析.结果:双酶切6 h,片段主要集中在250～2 000bp;连接产物和预扩增产物最适稀释倍数均为10倍;预扩增产物经选择性引物E-ACF/M-CAT和E-ACA/M-CAG扩增,琼脂糖电泳检测其主带分别集中在250～375 bp和500～750 bp,6%聚丙烯酰胺凝胶电泳检测及银染,条带清晰可辨.结论:该体系具有稳定性高、重复性好等优点,可用于华中五味子AFLP分析.     

吉富罗非鱼AFLP反应体系的建立与优化

本文以我国水产养殖主导品种吉富罗非鱼(farmed tilapia)为研究材料,进行扩增片段长度多态性(AFLP)反应体系的研究,初步建立了一套适合罗非鱼的AFLP反应体系。对体系中关键环节的优化结果如下:基因组DNA要求无降解和RNA污染,OD260/OD280比值在1.8~2.0之间;基因组DNA EcoRⅠ/MseⅠ37℃双酶切6h、20℃连接20h,连接产物10倍稀释,PCR预扩增产物稀释45倍作为选择性扩增的模板,选择性扩增产物经6%变性丙烯酰胺凝胶电泳检测可获得条带清晰、背景干扰小的图像。该体系的构建为今后进行罗非鱼群体遗传多样性分析、种质鉴定、重要经济性状分子标记的开发及遗传连锁图谱构建等方面研究提供了参考。     

华中五味子AFLP反应体系的建立

目的：建立一个适于华中五味子研究用的AFLP反应体系。方法：以华中五味子硅胶干燥嫩叶为试材,采用改良CTAB法提取到高质量DNA。通过琼脂糖凝胶电泳和聚丙烯酰胺凝胶电泳对MseⅠ/EcoRⅠ双酶切、连接、预扩增和选择性扩增过程中的关键因素进行分析。结果：双酶切6h,片段主要集中在250～2000bp;连接产物和预扩增产物最适稀释倍数均为10倍;预扩增产物经选择性引物E-ACT/M-CAT和E-ACA/M-CAG扩增,琼脂糖电泳检测其主带分别集中在250～375bp和500～750bp,6%聚丙烯酰胺凝胶电泳检测及银染,条带清晰可辨。结论：该体系具有稳定性高、重复性好等优点,可用于华中五味子AFLP分析。     

日本沼虾AFLP反应体系的建立

目的:建立一个适于日本沼虾研究用的 AFLP 反应体系.方法:以日本沼虾 DNA 为材料,对基因组酶切时间、选择性扩增中Mg2 、dNTP浓度、预扩增产物稀释倍数及选扩性引物M 3/E 3配比等进行了比较分析.结果:酶切5h,选扩 25ul PCR 反应体系中Mg2 2mmol/L,dNTP 1.2rmnol/L,预扩产物稀释 40 倍,选扩引物M 3/E 3配比为8:1,所得产物在毛细管电泳中可得剑稳定的结果.结论:该体系的构建为 AFLP 技术在日本沼虾相关研究中的应用奠定了基础.     

甘蔗双低频酶cDNA-AFLP体系的优化

目的：建立一个适合于甘蔗（Saccharum officenarum）为研究材料的cDNA-AFLP反应的优化体系。方法：用Rever-tAidTM First Strandc DNA Synthesis Kit反转录获得第一链,用Rnase H、E.coliDNA聚合酶Ⅰ和E.coliDNA连接酶合成双链cDNA,用双低频酶EcoRⅠ和PstⅠ对dscDNA酶切,连接,预扩增,和选择性扩增的关键因素进行分析。结果：500ng的dscDNA双酶切5h,将16℃过夜连接的产物稀释1倍用作预扩增的模板,预扩增产物稀释50倍作为选择性扩增模板,6%聚丙烯酰胺凝胶检测及银染,扩增条带均匀分布,清晰可辨且主要分布在200～2000bp之间。结论：该体系具有稳定性高,重复性好等优点,可用于甘蔗cDNA-AFLP分析。     

喜旱莲子草MSAP分析技术反应体系的建立

目的：建立一个适于研究喜旱莲子草DNA甲基化的MSAP分析体系。方法：以喜旱莲子草为材料,采用改良CTAB法提取基因组DNA,并对影响MSAP分析的关键步骤包括基因组DNA酶切反应时间、模板DNA连接产物稀释倍数、预扩增产物稀释倍数等进行了优化。结果：改良CTAB方法提取的DNA质量较好,酶切反应时间5h,酶切连接产物稀释10倍进行预扩增,预扩产物稀释10倍进行选择性扩增,所得PCR产物经6%聚丙烯酰胺电泳,条带清晰可辨,并能有效检测喜旱莲子草基因组DNA甲基化的程度和状态。结论：为研究喜旱莲子草表观遗传适应机制奠定了基础。     

小叶锦鸡儿基因组DNA的提取及AFLP反应体系的建立

以小叶锦鸡儿幼嫩叶片为材料,采用改良的SDS法提取其基因组DNA,通过优化AFLP技术体系中的几个主要因素,建立了适合小叶锦鸡儿的AFLP银染反应体系。改良的SDS法能通过在提取液中加入β-巯基乙醇防止氧化、加入PVP去除酚类等物质,获得满足AFLP分析要求的纯度高、完整性好的基因组DNA;用EcoRⅠ和MseⅠ37 ℃双酶切4 h后可以将500 ng的基因组DNA完全切开。酶切产物和接头经16℃连接过夜后,用带有1个选择性碱基的引物和带有3个选择性碱基的引物分别进行预扩增和选择性扩增,扩增产物经变性聚丙烯酰胺凝胶电泳分离,用AgNO₃染色,得到了清晰的指纹式样。小叶锦鸡儿AFLP反应体系的建立为利用该技术研究小叶锦鸡儿的遗传多样性奠定了实验基础。     

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.