人-鼠嵌合Fab片段的温度诱导表达

构建了具有λPRPL启动子高效表达人鼠嵌合Fab片段的温度诱导表达型载体pHZ01。并在大肠杆菌中表达了三种嵌合Fab片段：抗前列腺特异抗原(PSA)的嵌合Fab．抗溶菌酶(HEL)的嵌合Fab和抗破伤风类毒素(TT)的嵌合Fab．三种表达的可溶 性嵌合Fab都具有特异结合抗原的能力,嵌合Fab的CHI和CK区均为人源的。较之鼠源 Fab具有更好的应用前景。     

丙型肝炎病毒基因组C区和NS3区基因cDNA的融合及其在大肠杆菌中的表达与初步应用

通过逆转录（ＲＴ）－聚合酶链式反应（ＰＣＲ）,从中国人丙型肝炎病毒（ＨＣＶ）携带者的血清中扩增并克隆到２段ｃＤＮＡ片段,即ＨＣＶ基因组Ｃ区抗原基因Ｃ８３１ｃＤＮＡ片断（约５３０ｂｐ）和ＮＳ３区抗原基因Ｃ３３ｃｃＤＮＡ片段（约８６０ｂｐ）。Ｃ３３ｃｃＤＮＡ片段同Ｃ８３１ｃＤＮＡ片段经连接　　　肽Ｓｅｒ－Ｐｒｏ－Ｇｌｙ－Ｓｅｒ连接成为基因嵌合体Ｃ３３ｃ－Ｃ８３１（约１４００ｂｐ）。Ｃ３３ｃ－Ｃ８３１基因嵌合体同温控型原核表达载体ｐＢＶ２２０重组,构建成表达质粒ｐＢＶ／Ｃ３３ｃ－Ｃ８３１,并在大肠杆菌细胞中获得了重组嵌合抗原Ｃ３３ｃ－ＣＬ的表达。通过酶切分析和Ｗｅｓｔｅｒｎ免疫印迹法,对约占菌体可溶性蛋白９％的表达产物做了鉴定。采用ＴｒｉｔｏｎＸ－１００和盐析处理,获得粗提表达产物。粗提的表达产物经尿素裂解和离子交换层析纯化,得到可用于检测抗ＨＣＶ核壳蛋白和抗ＮＳ３区抗体的重组嵌合抗原Ｃ３３ｃ－ＣＬ。对Ｃ３３ｃ－ＣＬ做抗原性分析发现,它同时具有完整的Ｃ３３ｃ抗原和Ｃ２２抗原的免疫反应活性,完全能替代单纯的Ｃ３３ｃ和Ｃ２２抗原。该嵌合抗原在血清学诊断中有重要的应用价值,可望成为新一代ＨＣＶＥＩＡ诊断试剂的优选抗原。     

人抗体组合文库的构建和抗HBsAg噬菌体抗体的筛选

应用噬菌体表面递呈表达系统构筑建了人抗体组合文库,并同了结合乙肝表面抗原（ＨＢｓＡｇ）的人噬菌体抗体（Ｆａｂ片段）。以免疫球蛋白信号肽序列为引物进行半套式ＰＣＲ所得到的产物在质和量上优越于以可变区５末端保守库列为引物进行ＰＣＲ所得到的产物,经过３次亲和选择后,抗体阳性率为６９％,抑制实验表明,所筛选出来的噬菌体本具有抗ＨＢｓＡｇ的特异性,序更分析表明ＶＨ分别属于ＶＨＩ亚群和Ⅲ亚群,其轻链ＶＬ分别属     

丙型肝炎病毒抗原决定簇与霍乱毒素B亚基的融合表达以及融合蛋白的抗原性

通过固相化学合成法,合成了编码丙型肝炎病毒（ＨＣＶ）结构区和非结构区的４个抗原决定簇基因,这些抗原决定簇基因片段以不同方式串朕后与ｃｔｘＢ基因融合,构建了１２种表达不同嵌合蛋白的重组质粒。各重组质粒转化大肠杆菌后均能高效分泌性表达融合蛋白,表达产量随所融合的抗原决定簇不同在１０～５０μｇ／ｍｌ之间,表达水平主要与抗原决定簇的氨基酸组成有关,而与抗原决定簇的大小及串联次数关系不大。融合蛋白通过亲和层析纯化达到了电泳纯。对１１种融合蛋白中ＨＣＶ抗原决定簇的反应原性的研究表明,多数融合蛋白中ＨＣＶ抗原决定簇均能与对应的ＨＣＶ抗体结合。其中以融合蛋白９５０８２为抗原研制的抗－ＨＣＶＥＬＩＳＡ试剂具有良好的灵敏度和特异性。     

双专－性抗体介导的纤溶作用

用双功能团试剂将抗尿激酶单克隆抗体Ｎ３４的ＩｇＧ和抗人活化血小板α－颗粒膜蛋白ＧＭＰ－１４０单克隆抗体ＳＺ－５１的Ｆａｂ片段通过二硫键共价偶联,偶联的抗体保留了对各自抗原的亲和性。这种对尿激酶和血栓同时具有亲和活性的双专一性抗体（Ｎ３４－ＳＺ－５１）提高低分子量尿激酶的溶栓效率３８倍,且对血浆中纤维蛋白原的含量基本上不影响。     

双专一性抗体介导的纤溶作用

用双功能团试剂将抗尿激酶单克隆抗体Ｎ３４的ＩｇＧ和抗人活经血小板α－颗粒膜蛋白ＧＭＰ－１４０单克隆抗体ＳＺ－５１的Ｆ,ａｂ片段通过二硫键共价偶联,偶联的抗体保留了对各自抗原的亲和性。这种对尿再生产血栓同时具有亲和活性的双专一性抗体（Ｎ３４－ＳＺ－５１）提高低分子量尿激酶的溶栓效率３８倍,且对血浆中纤维蛋白原的含量基本上影响。     

基因工程重组嵌合抗原作为丙型肝炎诊断试剂的应用

利用基因工程重组技术获得了三种嵌合抗原,两种为丙型肝炎病毒（ＨＣＶ）结构区核壳蛋白Ｃ２２和ＮＳ３非结构区Ｃ３３ｃ双表位的嵌合抗原Ｂ１和Ｂ２,另一种是既具有上述两段优势抗原,还包括ＮＳ４非结构区Ｃ１００多表位的嵌合抗原Ｃ２５。在大肠杆菌水解酶阴性Ｂ株菌中表达的产物,经ＳＤＳ－ＰＡＧＥ分析,Ｂ１,Ｂ２和Ｃ２５嵌合抗原分子量分别为４３,５３和７０ｋＤ,抗原蛋白表达量分别各占电泳蛋白条带的４０％,１０％和     

人巨细胞病毒大外膜磷蛋白pp150羧基末端的高效表达及初步应用

以基因工程抗原取代天然抗原用于人巨细胞病毒（ＨＣＭＶ）感染的诊断,具有广阔的前景。为此,分离ＨＣＭＶ大外膜磷蛋白ｐｐ１５０基因ＯＲＦ３端４３４ｂｐ（编码羧基末端ａａ９４３～ａａ１０４８）的ＤＮＡ片段,导入带有高水平转录启动子Ｐｔｒｃ和六个串联组氨酸的原核表达载体ｐＴｒｃＨｉｓ,在大肠杆菌中得到高效表达,ＳＤＳ ＰＡＧＥ显示一明显的分子量为１５６ｋＤ的融合蛋白条带,ＬＫＢ激光扫描占菌体总蛋白的５１％以上,并且以可溶性形式存在。表达产物经高效特异性纯化后,ＥＬＩＳＡ结果表明其能够与ＨＣＭＶＩｇＭ阳性血清呈特异反应,为建立新一代ＨＣＭＶＩｇＭ检测试剂盒打下了基础。     

丙型肝炎病毒核心区优势抗原表位基因的化学合成及其表达产物的抗原性分析

依据丙型肝炎病毒（ＨＣＶ）多蛋白核心区Ｎ端氨基酸序列和密码子简并性,人为设计并采用半化学半酶促的方法合成了一个ＤＮＡ片段。经核酸杂交检测以及ＤＮＡ序列分析证实,该片段的核苷酸序列与设计完全一致。将合成的ＤＮＡ片段插入融合表达载体ｐＧＥＸ－２Ｔ中,表达产物经亲和层析纯化后进行Ｗｅｓｔｅｒｎ免疫印迹实验和间接ＥＬＩＳＡ分析,结果表明,融合蛋白具有ＨＣＶ核心区抗原的免疫反应性,可望用于ＨＣＶ抗体的检测．此外,编码ＨＣＶ优势抗原表位的化学合成基因有可能为ＨＣＶ嵌合抗原的研究提供一条捷径。     

通过构建轻链二级库对人源抗TNF-α抗体进行亲和力成熟

通过构建轻链二级库的方法,对人源抗ＴＮＦ－α单抗Ｆａｂ进行轻链置换,并筛选出具有更高亲和力的人源抗ＴＮＦ－α单抗的Ｆａｂ。首先用ＲＴ－ＰＣＲ技术扩增正常人全套抗体轻链基因,并与已获得的人源抗ＴＮＦ－α单抗的重链基因配对,构建人源抗ＴＮＦ－α噬菌体抗体轻链二级库,然后筛选与ＴＮＦ－α具有更高亲和力的克隆,经过三轮的生物淘筛（ｂｉｏｐａｎｎｉｎｇ）,获得了比原来的人源抗ＴＮＦ－α单抗Ｆａｂ具有更高亲和     

Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis

It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis.     

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.