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Methods Amplify the cDNA sequence encoding truncated HCV gene, with a part of carboxyl-terminus deleted, by PCR. Synthesize the mimic epitope at E2 region of HCV and seven T or Th cell epitope genes of NS3-NS5 respectively. Bind HCV core gene with a part of carboxyl-terminus deleted to the synthetic epitope gene by PCR, then clone into eukaryotic expression vector pcDNA3.1 and transiently transfect COS7 cells.

用PCR方法扩增核心区羧基端部分缺失的基因片段；分别合成HCV E2区模拟表位和NS3~NS5 7个T或Th细胞表位基因；PCR方法将羧基端部分缺失的HCV核心区基因与合成的表位基因串联，克隆入真核表达载体pcDNA3.1，并通过脂质体瞬时转染COS7细胞。

Rats living over 96h were considered survival. To observe the protective effects of ALR gene on acute liver injury rat, thirty-six rats that were injected peritoneally with 50% CCl4 of 2ml/kg were divided into the flowing 6 group according to doses (50?g/kg and 200?g/kg ) and administration routes of pcDNA3-ALR DNA 4h after CCl4 injection: group 1, model group; group 2, ALR gene of 50μg/kg was injected venally; group 3, ALR gene of 200?

另取36只大鼠按2ml/kg腹腔注射50% CCL4，染毒后4h按不同剂量（50μg/kg 和200μg/kg ）、不同注射途径（尾静脉注射、腹腔注射和联合注射）随机分为六组。1组：模型组，不注射ALR基因； 2组：尾静脉注射ALR基因 50μg/kg； 3组：尾静脉注射ALR基因 200μg/kg； 4组：腹腔注射ALR基因 50μg/kg； 5组：腹腔注射ALR基因 200μg/kg； 6组：联合注射（尾静脉和腹腔各注射ALR 基因100μg/kg）。

The invention discloses a pyruvate oxidase gene derived from viridian aerococcus, recombinant expression plasmid constructed by the gene and an engineering strain of escherichia coli gene transformed by the recombinant expression plasmid.

本发明公开了一种绿色气球菌来源的丙酮酸氧化酶基因AvPyOD，该基因所构建的重组表达质粒，以及该重组表达质粒转化的大肠杆菌基因工程菌株。

The method is simple, has safe operation, creates an approach utilizing a plant gene to safely select the marker gene for the tomato genetic transformation, can replace the disadvantages caused by the widely used heterogenous genes such as antibiotic genes, weedicide resistant genes, xyl A, pmi, GUS and so on, taken as marker genes, obtains a transgenic tomato with food safety, biological safety and ecological safety, relieves the public from anxiety of marker gene safety, and inevitably facilitates the commercialized application of the transgenic tomato.

该方法简单，操作安全，开创了利用植物基因作为番茄转化安全筛选标记基因的途径，能够代替目前广泛使用的抗生素基因、抗除草剂基因以及xylA、pmi、GUS等异源基因作为标记基因存在的弊端，获得具有食品安全性、生物安全性和生态安全性的转基因番茄，解除公众对标记基因安全性的顾虑，必将促进转基因番茄的商业化应用。

METHODS: MyoD cDNA fragments were extracted from plasmids pEMSV-MyoD with polymerase chain reaction, and PCR was used to clone the whole-length gene of MyoD. After adding CACC sequence at 5' end, MyoD gene was cloned by orient topology into transfer ventor, pENTR/D-TOPO. Objective gene was transferred into adenoviral expression vector DNA via pENTR/D-TOPO vector. The recombinant adenoviral vectors transfected into HEK293A cells by using lipofectamine were packaged and amplified.

从pEMSV-MyoD质粒上用聚合酶链反应法扩增出MyoD cDNA片段，再通过聚合酶链反应使MyoD基因加上CACC序列接头，经过定向拓扑克隆使目的基因连接到转移载体上，再通过LR酶促反应，将目的基因转移到腺病毒表达载体DNA上，获得MyoD基因重组的腺病毒DNA，用脂质体转染法转染HEK293A细胞，包装扩增出MyoD基因重组的腺病毒。

LacZ gene was fused with the promoter of the acoR gene and bkdR gene, and two recombined genes were expressed in sigL mutant strain and HD-73 wild strain sequentially.

分别将调控aco操纵子（编码3-羟基丁酮脱氢酶系统）的转录调节基因acoR和调控bkd操纵子（编码催化支链脂肪酸合成的酶系统）的转录调节基因bkdR的启动子与lacZ基因融合，并转入出发菌株和sigL突变体中，测定b-半乳糖苷酶的活性。

Chinensis,tissue culture,Hygromycin,genetic transformaction system,gene isolation,Agamous gene,brassinolide,cDNA array,responsive gene

中国水仙；组织培养；潮霉素；转化系统；基因克隆； Agamous基因；油菜素内酯； cDNA阵列；应答基因

The avian influenza virus RNA was directly extracted and purified from chicken em-bryo allantoic fluid. According to published gene sequence,a pair of specific primers to the nucleoprotein gene of AIV was designed and synthesized. After that,the NP cDNA was amplified by RT-PCR .Then the complete NP gene was sequenced.

直接从鸡胚尿囊液中提取H9N2亚型禽流感病毒的RNA，并根据已发表的A型流感病毒株的核苷酸序列，设计了1对特异引物，采用RT-PCR技术成功地扩增了AIV的NP基因；将NPcDNA克隆后进行了序列测定。

Molecular cloning and characterization of the novel lection gene. Primers were designed based on conserved sequences of other lectin genes from Amaryllidaceae. A novel lectin gene was isolated from Zephyrathes grandiflora. The full-length cDNA of Zephyrathes grandiflora lectin was 955bp, containing an open reading frame (576bp) encoding a peptide of 191 amino acids. The gene was named as zga (Zephyrathes grandiflora agglutinin), with its encoding protein showing 59% similarity to GNA.

基因克隆及序列分析：根据石蒜科植物凝集素的保守区段，设计引物，采用RACE技术从韭莲中克隆出长度为952bp的新凝集素基因，包含了完整的编码序列(576bp)，它编码一个全长191个氨基酸的蛋白，与雪花莲外源凝集的相似性达59％，被命名为ZGA(Zephyrathes grandiflora agglutinin)。

The stable clones are further identified by RT-PCR and Western blot; 6 MTT assay is used to investigate the effect of ZNRD1 on the cell growth of cells (AGS, SGC7901, MKN28, NIH3T3, GES-1); 7 Soft agar assay is used to investigate the effect of ZNRD1 on the clonality of cells (AGS, MKN28); 8 Nude mice assay is used to investigate the effect of ZNRD1 on the cell growth of gastric cancer cells (AGS, MKN28); 9 Flow cytometry is used to investigate the effect of ZNRD1 on the cell cycle distribution of cells (AGS, MKN28, NIH3T3, GES-1); 10 Flow cytometry is used to investigate the effect of ZNRD1 on the cell apoptosis of cells (AGS, MKN28, NIH3T3); 11 MTT assay is used to investigate the effect of ZNRD1 on the drug sensitivity of cancer cells (SGC7901, SGC7901/VCR, HL-60, HL-60/VCR) in vitro; 12 SRCA is used to investigate the effect of ZNRD1 on the drug sensitivity of gastric cancer cells (SGC7901, SGC7901/VCR) in vivo; 13 Flow cytometry is used to investigate the effect of ZNRD1 on adriamycin accumulation of cancer cells (SGC7901, SGC7901/VCR, HL-60, HL-60/VCR); 14 Transmission electron microscope is used to investigate the effect of ZNRD1 on the sensitivity of SGC7901 cells towards drug-induced apoptosis; 15 Flow cytometry and DNA ladder assay are used to investigate the effect of ZNRD1 on the sensitivity of cells (SGC7901, SGC7901/VCR, HL-60/VCR) towards drug-induced apoptosis; 16 Microarray is used to investigate the profiling of ZNRD1-responsive genes in gastric cancer cells (AGS, MKN28, SGC7901, SGC7901/VCR); 17 RT-PCR and Western blot are used to identify the results of microarray; 18 Reporter gene assay is used to investigate the effect of ZNRD1 on the transcriptional activity of cyclin D1; 19 Reporter gene assay is used to investigate the effect of ZNRD1 on the transcriptional activity of MDR1; 20 Kinase assay is used to investigate the effect of ZNRD1 on the activity of cyclin E-CDK2 kinase; 21 The antisensenucleic acids of p21 is used to inhibit the expression of p21, and flow cytometry is used to investigate the effect of p21 on ZNRD1-induced cell cycle arrest in gastric cancer cells; 22 The antisensenucleic acids of p27 is used to inhibit the expression of p27, and flow cytometry is used to investigate the effect of p27 on ZNRD1-induced cell cycle arrest in gastric cancer cells; 23 Liposome is used to up-regulate the expression of Skp2, and flow cytometry is used to investigate the effect of Skp2 on ZNRD1-induced cell cycle arrest in gastric cancer cells; 24 Western blot is used to investigate the effect of ZNRD1 on the stability of Skp2 and p27 in gastric cancer cells; 25 MVD assay is used to investigate the effect of ZNRD1 on the angiopoietic activity of gastric cancer cells; 26 ELISA is used to investigate the effect of ZNRD1 on the expression of VEGF165 in gastric cancer cells; 27 The roles of DARPP-32 in MDR of gastric cancer cells are investigated using gene transfection, MTT assay, SRCA, flow cytometry and DNA ladder assay.

应用杂交瘤技术制备ZNRD1的首个单克隆抗体；2）利用RT-PCR、Western blot和免疫组化检测ZNRD1在胃癌组织、胃炎组织、正常胃上皮组织、胃癌细胞和正常胃组织上皮细胞中的表达；3）构建ZNRD1的小干扰RNA载体，并测序鉴定；4）利用脂质体将ZNRD1的真核表达载体及其空载体转染胃癌细胞（AGS、SGC7901、MKN28）和小鼠成纤维细胞（NIH3T3），G418筛选后进行鉴定；5）利用脂质体将ZNRD1的小干扰RNA载体及其空载体转染药敏胃癌细胞（SGC7901）、正常胃组织上皮细胞（GES-1）、对长春新碱耐药的胃癌细胞（SGC7901/VCR）、药敏白血病细胞（HL-60）、对长春新碱耐药的白血病细胞（HL-60/VCR），G418筛选后进行鉴定；6）利用MTT实验检测ZNRD1高/低表达对细胞（AGS、SGC7901、MKN28、NIH3T3、GES-1）生长的影响；7）通过软琼脂克隆形成实验检测上调ZNRD1对AGS、MKN28细胞克隆形成能力的影响；8）通过裸鼠成瘤实验检测上调ZNRD1对AGS、MKN28细胞体内成瘤性的影响；9）通过流式细胞仪分析ZNRD1高/低表达对细胞（AGS、MKN28、NIH3T3、GES-1）的细胞周期的影响；10）通过流式细胞仪分析上调ZNRD1对细胞（AGS、MKN28、NIH3T3）的凋亡的影响；11）通过MTT实验检测ZNRD1高/低表达对细胞（SGC7901、SGC7901/VCR、HL-60、HL-60/VCR）体外药物敏感性的影响；12）通过肾包膜下移植法检测ZNRD1高/低表达对细胞（SGC7901、SGC7901/VCR）体内药物敏感性的影响；13）通过流式细胞仪分析ZNRD1高/低表达对细胞（SGC7901、SGC7901/VCR、HL-60、HL-60/VCR）内阿霉素蓄积和泵出的影响；14）通过透射电镜检测上调ZNRD1对SGC7901细胞凋亡敏感性的影响；15）通过流式细胞仪和DNA梯度试验检测ZNRD1高/低表达对细胞（SGC7901、SGC7901/VCR、HL-60）凋亡敏感性的影响；16）通过基因芯片检测ZNRD1高/低表达对胃癌细胞内基因表达谱的影响；17）利用RT-PCR、Western blot对基因芯片的结果进行鉴定；18）利用报告基因实验检测ZNRD1对cyclin D1的启动子活性的调节作用；19）利用报告基因实验检测ZNRD1高/低表达对MDR1的启动子活性的调节作用；20）利用激酶试验检测ZNRD1对cyclin E-CDK2 激酶活力的影响；21）利用反义核酸技术抑制p21的表达；通过流式细胞仪检测抑制p21对ZNRD1介导的细胞周期阻滞的影响；22）利用反义核酸技术抑制p27的表达；通过流式细胞仪检测抑制p27对ZNRD1介导的细胞周期阻滞的影响；23）利用脂质体转染法上调Skp2的表达；通过流式细胞仪检测上调Skp2对ZNRD1介导的细胞周期阻滞的影响；24）利用Western blot检测ZNRD1对p27和Skp2的蛋白稳定性的影响；25）利用微血管密度实验检测ZNRD1对AGS、MKN28细胞裸鼠移植瘤微血管形成的影响；26）利用ELISA检测ZNRD1对AGS、MKN28细胞培养上清和移植瘤匀浆中VEGF165含量的影响；27）利用脂质体转染法、MTT实验、肾包膜下移植法、流式细胞仪和DNA梯度试验检测新耐药相关分子DARPP-32对细胞（SGC7901、SGC7901/VCR、对阿霉素耐药的胃癌细胞SGC7901/ADR）多药耐药表型的影响；利用脂质体转染法和MTT实验检测下调ZNRD1对DARPP-32介导的胃癌多药耐药的调控作用。

The split between the two groups can hardly be papered over.

这两个团体间的分歧难以掩饰。

This approach not only encourages a greater number of responses, but minimizes the likelihood of stale groupthink.

这种做法不仅鼓励了更多的反应，而且减少跟风的可能性。