期刊文献+
共找到19篇文章
< 1 >
每页显示 20 50 100
TM9SF1 promotes bladder cancer cell growth and infiltration 被引量:2
1
作者 Long Wei Shi-Shuo Wang +9 位作者 Zhi-Guang Huang Rong-Quan He Jia-Yuan Luo Bin Li Ji-Wen Cheng Kun-Jun Wu Yu-Hong Zhou Shi Liu Sheng-Hua Li Gang Chen 《World Journal of Clinical Oncology》 2024年第2期302-316,共15页
BACKGROUND Bladder cancer(BC)is the most common urological tumor.It has a high recur-rence rate,displays tutor heterogeneity,and resists chemotherapy.Furthermore,the long-term survival rate of BC patients has remained... BACKGROUND Bladder cancer(BC)is the most common urological tumor.It has a high recur-rence rate,displays tutor heterogeneity,and resists chemotherapy.Furthermore,the long-term survival rate of BC patients has remained unchanged for decades,which seriously affects the quality of patient survival.To improve the survival rate and prognosis of BC patients,it is necessary to explore the molecular mechanisms of BC development and progression and identify targets for treatment and intervention.Transmembrane 9 superfamily member 1(TM9SF1),also known as MP70 and HMP70,is a member of a family of nine transmembrane superfamily proteins,which was first identified in 1997.TM9SF1 can be expressed in BC,but its biological function and mechanism in BC are not clear.AIM To investigate the biological function and mechanism of TM9SF1 in BC.Overexpression of TM9SF1 increased the in vitro proliferation,migration,and invasion of BC cells by promoting the entry of BC cells into the G2/M phase.Silencing of TM9SF1 inhibited in vitro proliferation,migration,and invasion of BC cells and blocked BC cells in the G1 phase.CONCLUSION TM9SF1 may be an oncogene in BC. 展开更多
关键词 TM9sf1 Bladder cancer Biological function cell function assay ONCOGENE
下载PDF
TM9SF1 is implicated in promoting the proliferation and invasion of bladder cancer cells
2
作者 Shu-Qing Zhou Lian-Xiang Luo 《World Journal of Clinical Oncology》 2024年第2期175-177,共3页
Zhuo et al looked into the part of transmembrane 9 superfamily member 1(TM9SF1)in bladder cancer(BC),and evaluated if it can be used as a therapeutic target.They created a permanent BC cell line and tested the effects... Zhuo et al looked into the part of transmembrane 9 superfamily member 1(TM9SF1)in bladder cancer(BC),and evaluated if it can be used as a therapeutic target.They created a permanent BC cell line and tested the effects of TM9SF1 overexpression and suppression on BC cell growth,movement,invasion,and cell cycle advancement.Their results show that TM9SF1 can boost the growth,movement,and invasion of BC cells and their access into the G2/M stage of the cell cycle.This research gives a novel direction and concept for targeted therapy of BC. 展开更多
关键词 Bladder cancer TM9sf1 cell proliferation Migration INVASION TM9sf1 overexpression TM9sf1 silencing inhibits
下载PDF
Human μ-opioid receptor overexpressed in Sf9 insect cells functionally coupled to endogenous G_(i/o) proteins 被引量:3
3
作者 WEI QIANG DE HE ZHOU +5 位作者 QING XIANG SHEN JIE CHEN LI WEI CHEN TIE LIN WANG GANG PEI ZHI QIANG CHI(e-mail:dhzhou@mail.shcnc.ac.cn)(1 Shanghai Institute of Materia Medica,2 Shanghai Institute of Cell Biology,Shanghai Academy of Life Sciences, Chinese Acad 《Cell Research》 SCIE CAS CSCD 2000年第2期93-102,共10页
Human μ-opioid receptor (HμOR) with a tag of six consecutive histidines at its carboxyl terminus had been expressed in recombinant baculovirus infected Sf9 insect cells.The maximal binding capacity for the [3H] di... Human μ-opioid receptor (HμOR) with a tag of six consecutive histidines at its carboxyl terminus had been expressed in recombinant baculovirus infected Sf9 insect cells.The maximal binding capacity for the [3H] diprenorphine and [3H]ohmefentanyl (Ohm) were 9.1± 0.7 and 6.52±0.23 nmol/g protein, respectively. The [3H] diprenorphine or [3H] Ohm binding to the receptor expressed in Sf9 cells was strongly inhibited by μ-selective agonists [D-Ala2], N-methylPhe4, glyol5]enkephalin (DAGO), Ohm, and morphine, but neither by δ nor by K selective agonist. Na+ (100 mM) and GTP (50 μM) could reduce HμOR agonists etorphine and Ohm affinity binding to the overexpressed HμOR. μ-selective agonists DAGO and Ohm effectively stimulated [35S]GTPγS binding (EC50 = 2.7nM and 6.9 nM) and inhibited forskolin- stimulated cAMP accumulation (IC50 = 0.9 nM and 0.3 nM). The agonist-dependent effects could be blocked by opioid antagonist naloxone or by pretreatment of cells with pertussis toxin (PTX). These results demonstrated that HμOR overexpressed in Sf9 insect cells functionally coupled to endogenous Gi/o proteins. 展开更多
关键词 Human μ-opioid receptor (HμOR) sf9 insect cells pertussis toxin (PTX) endogenous G_(i/o) proteins
下载PDF
Identification of TM9SF2 as a Candidate of the Cell Surface Marker Common to Breast Carcinoma Cells 被引量:1
4
作者 Samah Abou-Sharieha Yuh Sugii +4 位作者 Tuoya Dongwei Yu Ling Chen Heizou Tokutaka Masaharu Seno 《Chinese Journal of Clinical Oncology》 CSCD 2009年第1期1-9,共9页
OBJECTIVE We aimed identification of cell surface molecules, which might serve as diagnostic biomarkers or useful targets for therapies, in breast cancer. METHODS We developed unique DNA microarray coupled with spheri... OBJECTIVE We aimed identification of cell surface molecules, which might serve as diagnostic biomarkers or useful targets for therapies, in breast cancer. METHODS We developed unique DNA microarray coupled with spherical self-organizing map (sSOM) analysis to characterize cells and tissues by the cell surface markers. In the microarray 1,797 probes for human genes coding membrane bound proteins were spotted. With this microarray the gene expression profiles of eight breast carcinoma cell lines were compared to identify the genes that were commonly expressed in breast carcinomas but not in normal cells. RESULTS The gene expression profiles of sSOM from the eight breast carcinoma cell lines were successfully distinguished from that of normal breast tissue derived cells suggesting the presence of genes of interest, sSOMon the data extensively filtered revealed several candidate genes, of which expression was significant in carcinoma cells but low in normal cells. Finally, TM9SF2 was nominated through validations of PCR procedures together with CD24 and ErbB3, which are known breast carcinoma markers. TMgSF2 expression was further confirmed by immunological staining. Interestingly, TMgSF2 was found to be expressed in all the cell lines evaluated while CD24 and ErbB3 were not in all of the carcinoma cells, supporting their relationship in sSOM. Although physiological significance of TMgSF2 is unknown yet, siRNA treatment significantly inhibited the growth of MDA- MB-231 cells. CONCLUSION We propose TM9SF2 as a novel and useful diagnostic marker as well as a potential molecular target specific to breast carcinoma cells covering wide range of breast cancer. 展开更多
关键词 breast carcinoma cell surface marker spherical self-organizing map DNA microarray TM9sf2.
下载PDF
截短的丙型肝炎病毒核心蛋白基因和绿色荧光蛋白基因在Sf9细胞中的融合表达
5
作者 沈剑平 朱诗应 《临床检验杂志》 CAS CSCD 北大核心 2009年第4期267-269,共3页
目的构建含截短的HCV核心蛋白基因和绿色荧光蛋白基因的重组杆状病毒表达载体,探讨其在Sf9细胞中的表达和抗原性。方法用PCR法扩增截短的HCV核心蛋白基因片段(Ct)和增强型绿色荧光蛋白(EGFP)基因,将其插入转座子载体pFastBac1,构建成重... 目的构建含截短的HCV核心蛋白基因和绿色荧光蛋白基因的重组杆状病毒表达载体,探讨其在Sf9细胞中的表达和抗原性。方法用PCR法扩增截短的HCV核心蛋白基因片段(Ct)和增强型绿色荧光蛋白(EGFP)基因,将其插入转座子载体pFastBac1,构建成重组表达载体pFastCt-EGFP,转化大肠埃希菌DH10Bac,获得重组杆状病毒穿梭质粒BacmidCt-EGFP,转染昆虫Sf 9细胞。结果SDS-PAGE和western blot鉴定表明成功表达了分子量约40 000的融合蛋白。以ELISA法检测发现该融合蛋白能与28份抗HCV抗体阳性血清中的15份发生反应,阳性率为53.6%。结论该融合蛋白在昆虫Sf 9细胞中成功表达并具有一定的抗原性。 展开更多
关键词 丙型肝炎病毒 截短的核心蛋白 绿色荧光蛋白 昆虫sf9细胞
下载PDF
细粒棘球蚴Eg95-Eg.ferritin融合基因在昆虫细胞/杆状病毒中的表达 被引量:2
6
作者 李红兵 杨文 +6 位作者 陈健茂 刘强 王志辉 张昕 吴璟 郎多勇 王志昇 《宁夏医科大学学报》 2015年第2期120-124,F0004,共6页
目的利用昆虫细胞/杆状病毒系统表达细粒棘球蚴Eg95-Eg.ferritin融合蛋白,用于开发包虫病新型疫苗以及建立相关血清学诊断方法等研究。方法从细粒棘球蚴包囊中分离原头节,超声粉碎后提取总RNA为模板,通过RT-PCR扩增细粒棘球蚴Eg95和Eg.f... 目的利用昆虫细胞/杆状病毒系统表达细粒棘球蚴Eg95-Eg.ferritin融合蛋白,用于开发包虫病新型疫苗以及建立相关血清学诊断方法等研究。方法从细粒棘球蚴包囊中分离原头节,超声粉碎后提取总RNA为模板,通过RT-PCR扩增细粒棘球蚴Eg95和Eg.ferritin基因,采用基因拼接法将Eg95和Eg.ferritin融合,将该融合基因Eg95-Eg.ferritin插入到p Fast Bac DUAL载体中,构建重组转座载体后转化DH10Bac感受态细胞,获得重组Bacmid质粒后转染Sf-9昆虫细胞,传毒3代,对表达蛋白进行Western blot鉴定。结果成功克隆了Eg95和Eg.ferritin基因,通过柔性氨基酸linker成功获得了融合基因Eg95-Eg.ferritin,经PCR和酶切鉴定成功构建了重组质粒p Fast Bac DUAL-Eg95-Eg.ferritin,Western blot结果证实表达蛋白能够被包虫病人标准阳性血清识别。结论在Bac-to-Bac杆状病毒表达系统中成功表达了细粒棘球蚴Eg95-Eg.ferritin融合蛋白,与包虫病人标准阳性血清具有良好的反应性。 展开更多
关键词 细粒棘球蚴 Eg95-Eg.ferritin融合基因 杆状病毒 sf-9细胞 表达
下载PDF
Porcine Interleukin-2 Expression in Insect Cells and Its Enhancement of Pig Immunity to Swine Influenza Virus Inactivated Vaccine 被引量:3
7
作者 CHEN Hong-ying ZHANG Hong-ying HUANG Yan-quan CUI Bao-an WANG Zhen-ya WANG Yan-bin LIU Jin-peng CHAO An-jun 《Agricultural Sciences in China》 CAS CSCD 2010年第8期1211-1220,共10页
Mature porcine interleukin-2 (pIL-2) gene was amplified by PCR from the plasmid pGEM-T-pIL2 and cloned into the baculovirus pFastBacTM Dual vector of the Bac-to-Bac baculovirus expression system under the control of... Mature porcine interleukin-2 (pIL-2) gene was amplified by PCR from the plasmid pGEM-T-pIL2 and cloned into the baculovirus pFastBacTM Dual vector of the Bac-to-Bac baculovirus expression system under the control of the PH promoter. Recombinant plL-2 (rpIL-2) expressed in Sf9 insect cells was detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunofluorescence assay. Western blot analysis confirmed that the rpIL-2 protein had a molecular mass of 20 kDa, which was larger than the molecular mass of the mature protein predicted based on its peptide sequence. The rpIL-2 protein induced in vitro proliferation of ConA-stimulated porcine splenocytes and enhanced in vivo protective immune responses induced by vaccinating the pigs with inactivated oil emulsion vaccine against swine influenza virus. The results showed that the rpIL-2 expressed in Sf9 insect cells has immunoenhancement effects; the finding lays the foundation for the preparation of a specific recombinant IL-2 protein and the development of a novel immune adjuvant of vaccines against various infectious porcine pathogens to increase the immunoprotective efficacy of vaccines. 展开更多
关键词 porcine interleukin-2 sf9 insect cells EXPRESSION inactivated vaccine swine influenza virus
下载PDF
Expression of Rice Gall Dwarf Virus Outer Coat Protein Gene (S8) in Insect Cells
8
作者 Guo-cheng FAN Fang-luan GAO +5 位作者 Tai-yun WEI Mei-ying HUANG Li-yan XIE Zu-jian WU Qi-ying LIN Lian-hui XIE 《Virologica Sinica》 SCIE CAS CSCD 2010年第6期401-408,共8页
To obtain the P8 protein of Rice gall dwarf virus (RGDV) with biological activity,its outer coat protein gene S8 was expressed in Spodoptera frugiperda (Sf9) insect cells using the baculovirus expression system.The S8... To obtain the P8 protein of Rice gall dwarf virus (RGDV) with biological activity,its outer coat protein gene S8 was expressed in Spodoptera frugiperda (Sf9) insect cells using the baculovirus expression system.The S8 gene was subcloned into the pFastBacTM1 vector,to produce the recombinant baculovirus transfer vector pFB-S8.After transformation,pFB-S8 was introduced into the competent cells (E.coli DH10Bac) containing a shuttle vector,Bacmid,generating the recombinant bacmid rbpFB-S8.After being infected by recombinant baculovirus rvpFB-S8 at different multiplicities of infection,Sf9 cells were collected at different times and analyzed by SDS-PAGE,Western blotting and immunofluorescence microscopy.The expression level of the P8 protein was highest between 48-72 h after transfection of Sf9 cells.Immunofluorescence microscopy showed that P8 protein of RGDV formed punctate structures in the cytoplasm of Sf9 cells. 展开更多
关键词 Rice gall dwarf virus (RGDV) Outer coat protein Baculovirus expression system Spodoptera frugiperda sf9 insect cells
下载PDF
庚型肝炎病毒NS3蛋白在杆状病毒载体中的表达及其抗原性研究 被引量:3
9
作者 朱分禄 任浩 +4 位作者 朱诗应 谢南 潘卫 董辉 戚中田 《第二军医大学学报》 CAS CSCD 北大核心 2000年第9期849-852,共4页
目的 :利用 Bac- to- Bac HT杆状病毒系统在 Sf9昆虫细胞中表达庚型肝炎病毒 (HGV) NS3蛋白 ,并对表达产物的免疫原性进行研究。 方法 :将 HGV NS3基因片段定向克隆至转座载体 p Fast Bac HTa,转化 DH10 Bac感受态细胞 ,37℃振荡培养 4... 目的 :利用 Bac- to- Bac HT杆状病毒系统在 Sf9昆虫细胞中表达庚型肝炎病毒 (HGV) NS3蛋白 ,并对表达产物的免疫原性进行研究。 方法 :将 HGV NS3基因片段定向克隆至转座载体 p Fast Bac HTa,转化 DH10 Bac感受态细胞 ,37℃振荡培养 4h使发生转座 ,用抗生素平皿筛选重组 bacmid。脂质体介导转染 Sf9昆虫细胞 ,待细胞形态明显改变后收获细胞和培养上清液 ,将上清液再次感染 Sf9细胞以大量表达 HGV NS3重组蛋白。利用 SDS- PAGE和 Western- blotting方法分析重组蛋白。结果 :SDS- PAGE分析发现表达产物在 Mr43810处有一条明显的蛋白带 ,占细胞总蛋白量的 30 %以上 ;应用 Ni- NTA亲和层析柱获得了纯化的重组蛋白 ;Western- blotting显示该抗原可与 HGV RNA阳性血清发生特异反应。 结论 :获得了昆虫细胞内表达的 HGV NS3重组蛋白 ,并证明该蛋白有可能用于 HGV感染的抗体检测。 展开更多
关键词 免疫原性 杆状病毒载体 庚型肝炎 NS3蛋白
下载PDF
MPB64在杆状病毒系统中的表达纯化及免疫原性鉴定 被引量:1
10
作者 徐丹 姜潮 +3 位作者 陈昱 艾君 王晓艳 李校堃 《中国免疫学杂志》 CAS CSCD 北大核心 2014年第7期921-926,共6页
目的:在杆状病毒昆虫细胞表达系统中表达结核分枝杆菌蛋白MPB64,并鉴定其免疫原性。方法:目的基因MPB64连接到pFastBac载体,获得pFastBac-MPB64质粒转化DH10 Bac感受态,通过Tn7转座片段将目的基因转座到Bacmid中,得到Bacmid-MPB6... 目的:在杆状病毒昆虫细胞表达系统中表达结核分枝杆菌蛋白MPB64,并鉴定其免疫原性。方法:目的基因MPB64连接到pFastBac载体,获得pFastBac-MPB64质粒转化DH10 Bac感受态,通过Tn7转座片段将目的基因转座到Bacmid中,得到Bacmid-MPB64穿梭载体,脂质体包被后转染Sf 9细胞收获P1代病毒,重复转染Sf 9细胞三次获得高滴度的P4代病毒,收集细胞上清通过Q Sepharose FF和Ni亲和层析柱两步层析纯化得到目的蛋白MPB64。纯化的蛋白免疫BALB/c小鼠并检测血清中抗体滴度,ELISA检测MPB64蛋白刺激脾脏细胞产生IFN-γ的浓度,MTT法检测目的蛋白对免疫后小鼠脾脏细胞的增殖作用。结果:MPB64在昆虫细胞中成功表达,纯化后蛋白纯度达90%以上,蛋白产量为35 mg/L,纯化蛋白能有效刺激BALB/c小鼠产生抗体,提高小鼠脾脏细胞培养基中IFN-γ的含量,目的蛋白在0.2~100μg/ml之间对脾脏细胞有明显的促增殖作用。结论:成功在杆状病毒昆虫细胞表达系统中表达了具有免疫原性的MPB64蛋白,为生产结核病疫苗奠定了基础。 展开更多
关键词 MPB64 杆状病毒表达系统 sf 9细胞 结核分枝杆菌
下载PDF
HCV核心和截短的包膜蛋白E2基因在昆虫细胞中的表达及其抗原性 被引量:1
11
作者 朱诗应 赵平 +2 位作者 任浩 赵兰娟 戚中田 《第二军医大学学报》 CAS CSCD 北大核心 2004年第9期962-965,共4页
目的 :构建含 HCV全长核心基因和截短的包膜蛋白 E2基因的重组杆状病毒表达载体 ,研究其表达和抗原性。 方法 :以含 HCV全长 c DNA克隆的 p GEM- HCJ4质粒为模板 ,PCR扩增全长的 HCV核心抗原基因和截短的 E2基因片段 ,插入转座载体 p Fa... 目的 :构建含 HCV全长核心基因和截短的包膜蛋白 E2基因的重组杆状病毒表达载体 ,研究其表达和抗原性。 方法 :以含 HCV全长 c DNA克隆的 p GEM- HCJ4质粒为模板 ,PCR扩增全长的 HCV核心抗原基因和截短的 E2基因片段 ,插入转座载体 p Fast Bac HTa构建重组转座质粒 p FB- CE2 t,转化 DH10 Bac大肠杆菌 ,获得重组杆状病毒穿梭质粒 Bacmid- CE2 t,以之转染昆虫 Sf9细胞进行外源基因的表达 ,并对其抗原性进行 EL ISA检测。 结果 :SDS- PAGE和 Western blot分析表明Bacmid- CE2 t在 Sf9细胞表达了 HCV C- E2蛋白 ,并能利用细胞内蛋白酶将其裂解为单独的 C蛋白和截短的 E2蛋白。纯化后的蛋白能分别与 HCV C蛋白、E2蛋白单抗以及慢性丙型肝炎患者血清反应。 结论 :HCV核心蛋白和截短的 展开更多
关键词 肝炎病毒 丙型 核心蛋白 截短的E2蛋白 昆虫sf 9细胞
下载PDF
重组截短型人角质细胞生长因子1在昆虫细胞中的表达 被引量:1
12
作者 薛萍 朱小静 +4 位作者 刘孝菊 李一兰 南佳 姜潮 李校堃 《吉林大学学报(医学版)》 CAS CSCD 北大核心 2012年第4期633-639,共7页
目的:探讨重组截短型人角质细胞生长因子1(rhKGF1D23)蛋白在昆虫细胞中表达的可行性,为rhKGF1D23蛋白的生产提供新的途径。方法:PCR法扩增昆虫偏爱密码子优化截短序列kgf1d23亚克隆到pFastbac载体。在辅助质粒的作用下,pFastBac-kgf1d2... 目的:探讨重组截短型人角质细胞生长因子1(rhKGF1D23)蛋白在昆虫细胞中表达的可行性,为rhKGF1D23蛋白的生产提供新的途径。方法:PCR法扩增昆虫偏爱密码子优化截短序列kgf1d23亚克隆到pFastbac载体。在辅助质粒的作用下,pFastBac-kgf1d23载体Tn7片段转座到Bacmid中mini-attTn7位点,得到穿梭载体Bacmid-kgf1d23,并通过脂质体法转染sf-9细胞得到杆状病毒P1。继续转染细胞扩增病毒并提高病毒滴度,蛋白表达分析从P3代病毒转染细胞开始,收集细胞超声裂解后进行纯化,MTT法检测纯化后目的蛋白生物活性。结果:昆虫偏好密码子改造的kgf1d23基因构建进转移载体pFastBac-kgf1d23和穿梭载体Bacmid-kgf1d23载体中。SDS-PAGE结果表明,重组蛋白在60h后开始大量表达,优化的收获蛋白时间在84~96h;利用肝素亲和层析和SP阳离子交换层析可以去除90%以上的杂蛋白。纯化的rhKGF1D23蛋白在0.3~25.0μg.L-1时,随浓度的增加HaCat细胞的促有丝分裂能力增强,在25.0μg.L-1时达到平台期。结论:重组截短型角质细胞生长因子rhKGF1D23蛋白在昆虫细胞中得到表达,保持其特有的肝素亲和特性,并具有免疫原性及细胞生物学活性。 展开更多
关键词 角质细胞生长因子1 截短突变 sf-9细胞 杆状病毒系统
下载PDF
草地贪夜蛾(Spodoptera frugiperda)细胞在生物反应器中培养条件初探
13
作者 余泽华 姚汉超 +3 位作者 陈曲侯 庞义 邓日强 龙綮新 《华中师范大学学报(自然科学版)》 CAS CSCD 北大核心 1999年第3期408-411,共4页
对草地贪夜蛾( Spodoptera frugiperda) Sf9 细胞在2 L Biostat R M D 型搅拌式生物反应器中进行了培养,并对细胞的生长状况、葡萄糖的消耗等进行了测定;对重组病毒的增殖、外源基因的... 对草地贪夜蛾( Spodoptera frugiperda) Sf9 细胞在2 L Biostat R M D 型搅拌式生物反应器中进行了培养,并对细胞的生长状况、葡萄糖的消耗等进行了测定;对重组病毒的增殖、外源基因的表达进行了初步探索.结果表明:细胞在该生物反应器中生长良好,细胞群体倍增时间33.4 h.病毒感染率大约在 89.8% 以上。 展开更多
关键词 sf-9昆虫细胞 生物反应器 细胞培养 草地贪夜蛾
下载PDF
嵌合卵清蛋白-兔出血症VP60蛋白病毒样颗粒在杆状病毒表达系统中的表达及其鉴定 被引量:1
14
作者 谭晖 胡波 +4 位作者 陈萌萌 范志宇 魏后军 宋艳华 王芳 《江苏农业学报》 CSCD 北大核心 2013年第2期352-358,共7页
利用卵清蛋白(OVA)第257~264位氨基酸替换兔出血症病毒的主要结构蛋白质VP60不同位置处相应数量的氨基酸,研究了氨基酸的替换对VP60蛋白表达及病毒样颗粒装配的影响。通过重叠延伸剪接术(SOE)法,OVA T-细胞表位(257~264位氨基酸)分别... 利用卵清蛋白(OVA)第257~264位氨基酸替换兔出血症病毒的主要结构蛋白质VP60不同位置处相应数量的氨基酸,研究了氨基酸的替换对VP60蛋白表达及病毒样颗粒装配的影响。通过重叠延伸剪接术(SOE)法,OVA T-细胞表位(257~264位氨基酸)分别取代VP60 3个区域(502~510位氨基酸、411~417位氨基酸、302~309位氨基酸),得到3种重组嵌合VP60基因。利用杆状病毒表达系统,得到3种重组穿梭载体Bacmid,分别命名为Bacmid-V-1、Bacmid-V-2、Bacmid-V-3。将重组阳性质粒转染Sf9昆虫细胞,得到3种重组病毒rBac-v-1、rBac-v-2、rBac-v-3,3组重组病毒感染Sf9细胞后表达的3种蛋白质分别命名为Z1、Z2和Z3。经RT-PCR、IFA、SDS-PAGE、Western blot等方法鉴定后,利用电镜观察此3种重组蛋白质病毒样颗粒的形成情况。3种重组嵌合VP60蛋白在Sf9昆虫细胞中得到表达,3种重组嵌合蛋白质均可以形成与VP60蛋白类似的病毒样颗粒。 展开更多
关键词 兔出血症病毒(RHDV) VP60蛋白 卵清蛋白 sf9细胞 病毒样颗粒(VLPs)
下载PDF
草地贪夜蛾细胞株Sf 9生测杀虫剂毒力的方法探索 被引量:1
15
作者 李春娜 张伟琪 +2 位作者 任豆豆 董兆克 张志勇 《应用昆虫学报》 CAS CSCD 北大核心 2020年第6期1279-1286,共8页
【目的】为适应杀虫剂新产品研发和害虫抗药性监测对快速生物测定的需要,本文探索细胞生测与试虫生测的关联情况。【方法】分别测定草地贪夜蛾Spodoptera frugiperda卵巢细胞株(Sf 9细胞)和小菜蛾Plutellaxyllostella敏感品系对不同类... 【目的】为适应杀虫剂新产品研发和害虫抗药性监测对快速生物测定的需要,本文探索细胞生测与试虫生测的关联情况。【方法】分别测定草地贪夜蛾Spodoptera frugiperda卵巢细胞株(Sf 9细胞)和小菜蛾Plutellaxyllostella敏感品系对不同类型的杀虫剂的毒力,并在此基础上进行了细胞活力抑制率与活体昆虫致死率的关联性分析。【结果】3种不同机理杀虫剂毒力在2种生测体系中均表现出阿维菌素>吡虫啉>高效氯氰菊酯的一致规律;同一类型(有机磷类)5种杀虫剂毒力在2种生测体系中均表现出马拉硫磷>敌敌畏>辛硫磷>丙硫磷>敌百虫的一致规律;同一种类(5%阿维菌素)3种剂型杀虫剂毒力在2种生测体系中均表现出了微乳剂>乳油>水乳剂的一致规律。细胞生测和试虫生测之间存在高度相关性,据此建立不同杀虫剂细胞8 h生测和试虫48 h生测剂量之间的线性回归方程。【结论】建立了不同杀虫剂处理Sf9细胞8h与处理小菜蛾幼虫48h的线性相关模型,可根据细胞生物测定结果反映试虫生物测定的结果,为今后新型农药的筛选、毒力测定等提供便利的条件。 展开更多
关键词 sf 9细胞 小菜蛾 杀虫剂 生物测定 LC50
原文传递
重组杆状病毒感染悬浮昆虫细胞及重组蛋白表达策略的优化
16
作者 李文丽 刘霞 +2 位作者 刘在新 卢曾军 范朋举 《河南农业科学》 CSCD 北大核心 2018年第8期118-122,共5页
为确定嵌合口蹄疫病毒(FMDV)中和表位的猪细小病毒(PPV)衣壳蛋白在昆虫细胞(Sf-9)中的最佳表达条件。通过对比悬浮培养和贴壁培养2种培养条件,探究Sf-9细胞的最佳培养方式;利用血球计数板每隔24 h进行细胞计数,绘制Sf-9细胞生长曲线;将P... 为确定嵌合口蹄疫病毒(FMDV)中和表位的猪细小病毒(PPV)衣壳蛋白在昆虫细胞(Sf-9)中的最佳表达条件。通过对比悬浮培养和贴壁培养2种培养条件,探究Sf-9细胞的最佳培养方式;利用血球计数板每隔24 h进行细胞计数,绘制Sf-9细胞生长曲线;将P1代重组病毒传代至P3代,提取重组病毒基因组DNA,进行PCR鉴定,检测插入目的基因的完整性;通过研究感染复数(MOI)和感染时间的关系来摸索重组病毒最佳感染条件,以获得高滴度的重组病毒;利用Western blot检测目的蛋白的表达量并探究重组蛋白的最佳表达条件。结果表明,悬浮培养的Sf-9细胞大小均一、边缘整齐,生长状态良好;PCR鉴定结果表明,插入的FMDV中和表位基因完整,可以进行重组蛋白的表达;Western blot结果显示,重组蛋白同时与FMDV阳性血清和PPV阳性血清产生特异性反应,进一步证明了插入的FMDV中和表位的存在;重组蛋白最佳表达条件是以10个MOI病毒液感染悬浮Sf-9细胞,在72 h时重组蛋白表达量最大。 展开更多
关键词 口蹄疫病毒 猪细小病毒 中和表位 杆状病毒载体表达系统 昆虫细胞
下载PDF
禽流感病毒H5N1亚型HA蛋白在昆虫细胞中的表达、纯化及反应原性分析 被引量:2
17
作者 仝晓丹 王玉娟 +6 位作者 杨仕标 盛东北 赵文华 黄晓静 谭菲菲 王同燕 田克恭 《黑龙江畜牧兽医》 CAS 北大核心 2021年第10期72-76,共5页
为了获得可溶性的禽流感病毒H5N1亚型HA蛋白,并将其用于单克隆抗体的制备和筛选,试验利用Bac-to-Bac杆状病毒表达系统进行HA蛋白的表达和纯化,以含全长HA基因的重组质粒为模板,经PCR扩增得到HA片段并克隆至pFastBac1载体,将鉴定正确的... 为了获得可溶性的禽流感病毒H5N1亚型HA蛋白,并将其用于单克隆抗体的制备和筛选,试验利用Bac-to-Bac杆状病毒表达系统进行HA蛋白的表达和纯化,以含全长HA基因的重组质粒为模板,经PCR扩增得到HA片段并克隆至pFastBac1载体,将鉴定正确的供体质粒转化至DH10Bac感受态细胞,获得Bacmid重组质粒,转染Sf 9细胞后获得重组杆状病毒。通过Western-blot和间接ELISA验证HA蛋白的表达情况和反应原性。结果表明:成功获得含有禽流感病毒H5N1亚型HA基因重组杆状病毒,HA蛋白可以分泌表达,大小约为63 ku,与预期大小一致,且能与H5N1标准阳性血清发生特异性反应;纯化的HA蛋白与免疫小鼠血清反应性良好。说明本研究成功表达并纯化获得了禽流感病毒H5N1亚型HA蛋白,并且具有良好的反应原性。 展开更多
关键词 禽流感病毒 血凝素蛋白 杆状病毒表达系统 蛋白纯化 免疫印迹分析 反应原性分析 sf 9细胞
下载PDF
Cytotoxicity and binding profiles of activated CrylAc and Cry2Ab to three insect cell lines 被引量:3
18
作者 Jizhen Wei Gemei Liang +4 位作者 Kongming Wu Shaohua Gu Yuyuan Guo Xinzhi Ni Xianchun Li 《Insect Science》 SCIE CAS CSCD 2018年第4期655-666,共12页
While CrylAc has been known to bind with larval midgut proteins cad- herin, APN (amino peptidase N), ALP (alkaline phosphatase) and ABCC2 (adenosine triphosphate-binding cassette transporter subfamily C2), littl... While CrylAc has been known to bind with larval midgut proteins cad- herin, APN (amino peptidase N), ALP (alkaline phosphatase) and ABCC2 (adenosine triphosphate-binding cassette transporter subfamily C2), little is known about the recep- tors of Cry2Ab. To provide a clue to the receptors of Cry2Ab, we tested the baseline cytotoxicity of activated Cry 1Ac and Cry2Ab against the midgut and fat body cell lines of Helicoverpa zea and the ovary cell line ofSpodopterafrugiperda (SFg). As expected, the descending order of cytotoxicity of CrylAc against the three cell lines in terms of 50% lethal concetration (LC50) was midgut (31.0μg/mL) 〉 fat body (59.0μg/mL) and SF9 cell (99.6μg/mL). By contrast, the fat body cell line (LC50 = 7.55μg/mL) was about twice more susceptible to Cry2Ab than the midgut cell line (16.0/xg/mL), the susceptibility of which was not significantly greater than that of SF9 cells (27.0μg/mL). Further, ligand blot showed the binding differences between CrylAc and Cry2Ab in the three cell lines. These results indicated that the receptors of Cry2Ab were enriched in fat body cells and thus largely different from the receptors of CrylAc, which were enriched in midgut cells. 展开更多
关键词 Bt cell toxicity RECEPTORS midgut cell fat body cell sf9 cell
原文传递
Molecular Cloning and Expression of Human Interleukin-6 in Insect Cells
19
作者 赵春文 王嘉玺 +1 位作者 肖定华 马贤凯 《Science China Chemistry》 SCIE EI CAS 1994年第9期1073-1081,共9页
670-bp hIL-6 cDNA fragments have been amplified by polymerase chain reaction(PCR)using recombinant plasmid pBMIL-6A as templates and two synthetic oligonucleotides containing the opti-mired translation initiation sequ... 670-bp hIL-6 cDNA fragments have been amplified by polymerase chain reaction(PCR)using recombinant plasmid pBMIL-6A as templates and two synthetic oligonucleotides containing the opti-mired translation initiation sequence/and restriction sites suitable for cloning as primers.The amplified IL-6cDNA fragments have then been recombined with a non-fusion expression baculovirus vector pVL1393.Theresultant recombinant plasmid pVL.IL-6 together with wtAcMNPV DNAs were transferred into culturedlepidopteran insect cells(Sf9)by calcium phosphate coprecipitation procedure.The recombinant baculovirus-es were formed by homologous recombination in vivo between pVL.IL-6 and wtAcMNPV DNAs,screenedfor plaque assay,and identified by means of dot blotting hybridization.The expressed rhIL-6 was secretedinto the culture medium,and its bioactivity was measured through half-maximum H-TdR incorporation intoIL-6-dependent cells 7TD1.As a result,the supernatant collected from recombinant baculovirus rAc.IL-6-infected Sf9 cells showed IL-6 activity of 10~6U/mL.The expression level of rhIL-6 of the supernatant deter-mined by IL-6 ELISA quantitation kit was 1 μg/mL. 展开更多
关键词 hIL-6 PCR BACULOVIRUS VECTOR wtAcMNPV sf9 cellS
原文传递
上一页 1 下一页 到第
使用帮助 返回顶部