光对商陆发状根生长和PAP基因表达的影响

利用发根农杆菌菌株Ar1334与美洲商陆(Phytolacca americana)叶片外植体共培养转化体系,共获得58个发状根无性系(SL-1~58).以发根农杆菌Ri质粒TL-DNA上的rol C基因设计特异引物,对发状根进行PCR检测,得到了预期的560 bp目的片段,表明Ri质粒T-DNA整合到发状根基因组中.将筛选出的株系SL-7接种在MS培养基上分别置于光、暗条件下进行培养.结果发现:SL-7在暗培养条件下呈乳白色,具有多分枝、多根毛、无向地性等典型的发状根特性;在光培养条件下,发根呈粉红色,少分支且生长缓慢;以商陆抗病毒蛋白(pokeweed antiviral protein,PAP)cDNA片段为探针,分别对光、暗条件下的发状根进行Northern blot检测,发现光对PAP基因的转录具有一定抑制作用;将发状根粗蛋白提取液与TMV病毒液混合后,摩擦接种于心叶烟(Nicotiana glutinosa)离体叶片,发现暗培养的发状根粗蛋白提取液对TMV抗性明显提高.表明商陆发状根的生长及PAP基因的表达都受到光的负向调控.该结果为商陆发状根的规模化培养和PAP蛋白的离体合成优化体系的建立奠定了基础.     

除虫菊发状根的诱导及培养条件优化

以除虫菊（Pyrethrum cinerariifolium Trey.）无菌苗为外植体,研究除虫菊发状根的诱导、培养条件优化,并对发状根中的除虫菊素进行检测和生物活性测定。结果显示,乙酰丁香酮能促进除虫菊下胚轴和子叶发状根的诱导,当乙酰丁香酮浓度为150 μmol/L时除虫菊下胚轴和子叶的诱导率为对照的2.29倍和2.66倍,预培养6 d时,下胚轴发状根诱导率为对照的2.25倍,发根农杆菌A4的诱导率均高于ATCC15834,愈伤组织较适合发状根的诱导,愈伤组织侵染后适合在无激素的MS培养基上进行发状根诱导,250 mL三角瓶中添加50 mL MS培养基较适合发状根的生长。对除虫菊发状根进行PCR检测发现,发根农杆菌含有的Ri T-DNA的rolB基因已整合进入发状根基因组中。通过GCMS检测发现,愈伤组织中除虫菊素的6种成分均未检测到,而发状根中检测到瓜菊素Ⅰ、茉酮菊素Ⅰ和茉酮菊素Ⅱ 3种成分,发状根对粘虫的拒食作用明显优于愈伤组织。本研究为通过组织培养方式生产除虫菊素奠定了基础。     

发根农杆菌诱导桑树毛状根体系的建立

应用发根农杆菌ACCC10060,以直接接种和共培养2种方法侵染桑树10 d龄子叶,并将2种处理的外植体分别接种于MS+AS(乙酰丁香酮,100 μmol/L)的平板,暗培养2 d后转接至MS+CS(头孢霉素,200 mg/L)平板培养3周,每3 d转接1次以除去其中所含的发根农杆菌菌体,结果2种侵染方法均成功诱导桑树产生毛状根,诱导效率分别为14%和17%.在无激素MS培养基上离体培养除菌后的毛状根,呈现旺盛的生长态势和典型的发状根结构特点.CTAB法提取毛状根基因组并进行PCR检测,结果扩增出了423 bp的rolB基因片段,表明Ri质粒的T-DNA已经成功整合到桑树的基因组中.     

农杆菌转化的小冠花发状根的诱导及其植株再生

利用野生型发根农杆菌15834菌株感染小冠花15日龄无菌苗子叶和下胚轴切段,建立了高效的发状根培养及其体细胞胚胎发生再生体系。发状根可直接从受伤的外植体表面产生,也能在外植体诱导的愈伤组织上发生,在无外源激素的MS固体和液体培养基上,转化根能自主生长,表现出典型的发根特征。用适宜浓度的乙酰丁香酮处理对数生长期的农杆菌菌液2h,感染预培养2d的子叶获得了最高的转化频率(87.4%)。在附加0.2mgL2,4_D,0.5mgLNAA和0.5mgLKT的MS培养基上,发状根能100%形成胚性愈伤组织,并于含0.5mgLKT,0.2mgLIBA和300mgL脯氨酸的MS培养基上顺序经过体细胞胚胎发育的各个典型时期,转换成完整植株。再生植株除具有发达的侧根外,其它形态特征与未转化植株未见明显的差异,但在获得的5个转化克隆中,其中1个的发状根及其再生植株叶片中有毒物质3_硝基丙酸的含量显著下降,分别为未转化对照的57.68%和58.17%。冠瘿碱纸电泳检测和rolB基因PCR扩增检测均证明农杆菌Ri质粒上的T_DNA已经整合到小冠花转化细胞的基因组中。     

新疆紫草毛状根的诱导及培养

将处于对数生长期(A600为0.5)的发根农杆菌MSU440、A4、R1000、15834、1025和R1601与新疆紫草子叶外植体共培养.结果表明:(1)发根农杆菌不同菌种对转化率有显著影响,供试6个菌种中只有MSU440菌株获得转化株.PCR及序列分析表明发根农杆菌Ri质粒的rolC基因已在新疆紫草毛状根基因组中整合并得到表达,转化率达4.5%.(2)子叶较真叶的不定根发生率高,且生根持续时间长.(3)在B5无铵无激素固体培养基上,毛状根分支多且根较长,达2~3 cm,毛状根鲜重月平均增殖达7~9倍,是固体培养毛状根的适宜培养基.(4)毛状根在MS无铵无激素液体培养基中培养12 d时,毛状根鲜重平均增殖达12倍,MS无铵液体培养基有利于毛状根的扩大生产.首次获得了激素自主、快速伸长生长、多分支、多根毛的新疆紫草毛状根株系,初步建立了新疆紫草毛状根诱导体系,为大规模培养、生产紫草素奠定了基础.     

木豆毛状根的诱导及其培养条件的研究

利用发根农杆菌LBA9402对木豆叶片直接进行诱导产生毛状根。本实验研究出诱导木豆毛状根的最佳条件是,以木豆叶片为外植体,于1/2MS固体培养基上预培养2~4 d,菌液浓度OD₆₀₀=0.6~0.8,浸染20 min,共培养3 d,诱导率为60.00%。在分子水平用PCR检测表明,发根农杆菌9402Ri质粒上的T-DNA成功整合进木豆毛状根的基因组中。     

4种发根农杆菌对朱砂根组培无菌叶片毛状根诱导的影响

以朱砂根(Ardisia crenata Sims)组培无菌叶片为材料,用4种发根农杆菌菌株(A4、ATCC15834、LBA9402和R1601)分别侵染进行毛状根诱导,比较朱砂根叶片毛状根诱导的最适培养基种类、预培养时间、侵染方式、共培养时间以及不同发根农杆菌的致根能力。研究表明:(1)朱砂根无菌叶片毛状根诱导最适培养基为1/2MS培养基,预培养2d、共培养2d,毛状根诱导率最高(31.87%)。(2)最佳侵染方式以剪好的幼叶和活化好的菌液(100mg/L AS)一起在28℃、180r/min黑暗条件下共振荡8~15min。(3)4种发根农杆菌均能诱导朱砂根叶片毛状根产生,但A4、ATCC15834效果最好,其致根能力大小顺序依次为ATCC15834A4LBA9402R1601。(4)PCR分子鉴定表明,发根农杆菌Ri质粒T-DNA已成功整合到宿主细胞核基因组中。     

宁夏枸杞发根农杆菌转化系的建立及影响转化因素的研究

以发根农杆菌Ａ４菌株介导,对宁夏枸杞叶片和茎切段的遗传转化进行了初步研究,建立了发状根体系,并优化了转化条件。乙酰丁香酮的添加、农杆菌液的浓度、共培养时间、外植体取材部位及时间,均可以影响发状根的诱导频率。采用添加１００μｍｏｌ／Ｌ的 乙酰丁香酮、振荡培养２４ｈ的Ａ４菌液感染３周龄的叶片切段,并共培养３ｄ,可以得到最佳的转化效果,发状根的诱导率为４８．９％。在同样的转化条件下,只有１３．６％的茎切     

发根土壤杆菌体外转化甘草子叶及下胚轴

利用发根土壤杆菌农杆碱型15834、A_4菌株和甘露碱型8196、K_(599)菌株体外转化甘草子叶及下胚轴。结果表明:除K_(599)菌株外,其余三种菌株都能诱导产生甘草发状根无性系。其中15834菌株的诱导率最高,下胚轴的诱导率高于子叶。组织学观察表明,在感染后4天左右,下胚轴维管束鞘外细胞、特别是形成层部分细胞大量启动形成分生细胞及分生细胞团,并进一步分裂形成根原基。6—8天后根原基具单向极性生长形成完整的发状根结构。发状根能在无外源激素的培养基上迅速生长。高压纸电泳检测到发状根中存在农杆碱及甘露碱,说明Ri质粒T-DNA编码的这两种冠瘿碱合成酶基因在甘草细胞中得到了表达。     

荞麦高频离体再生及发根农杆菌转化体系的建立

荞麦无菌苗下胚轴切段在不同激素配比的MS培养基上诱导愈伤组织,出愈率均为100％。在2．0mg/L2,4-D和1．5mg/L 6-BA组合下诱导产生的愈伤组织;转入2．0mg/L 6-BA和1．0mg/L KT的MS培养基,再生苗分化率在80％以上。根尖色体分析表明再生植株具一定的遗传稳定性。发根农杆菌A4转化荞麦下胚轴和子叶获得发状根,纸电泳检测所有随机取样测定的发状根均有相应冠瘿碱的存在。     

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.