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基于简并引物同步克隆两个红麻PDIL同源基因cDNA 5'-末端序列 被引量:2
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作者 金刚 陈鹏 +1 位作者 唐向民 周瑞阳 《生物技术通报》 CAS CSCD 北大核心 2011年第10期109-113,共5页
探索利用同一套简并引物结合通用引物同步扩增两个红麻PDIL同源基因的cDNA 5'-末端序列,以期为同一转录组中两个旁系同源基因cDNA 5'RACE的同步扩增提供借鉴。通过红麻HcPDIL5-2a和HcPDIL5-2b cDNA中间片段及3'-末端已知序... 探索利用同一套简并引物结合通用引物同步扩增两个红麻PDIL同源基因的cDNA 5'-末端序列,以期为同一转录组中两个旁系同源基因cDNA 5'RACE的同步扩增提供借鉴。通过红麻HcPDIL5-2a和HcPDIL5-2b cDNA中间片段及3'-末端已知序列的比对,在其完全保守区段设计了一条引物用于两个基因5'RACE的共反转录;在其部分保守区段设计了两条简并引物,并利用其在两个基因的5'RACE扩增时退火温度的差异,结合通用引物巢式PCR同步扩增两个基因的cDNA 5'-末端未知序列。在两个基因全长cDNA拼接序列的基础上设计两对特异引物分别扩增它们的cDNA全长序列,测序结果进一步验证了序列拼接和cDNA 5'RACE同步扩增的可靠性。进化分析证实两个基因属于PDIL基因家族成员。 展开更多
关键词 红麻 简并引物 5'RACE 同步克隆
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多克隆同步筛选及验证胰腺癌肺转移细胞亚群 被引量:1
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作者 田文嘉 杜洪震 +1 位作者 卞晓翠 李开龙 《中国老年学杂志》 CAS CSCD 北大核心 2012年第10期2122-2123,共2页
目的利用混合多克隆细胞亚群筛选和验证分析具有肺转移能力细胞亚群和基因。方法混合多克隆细胞亚群,分别尾静脉接种于喂饮强力霉素和对照两组裸小鼠,取小鼠肺原代培养得到具有肺转移能力的胰腺癌细胞亚群;提取基因组DNA进行PCR分析,比... 目的利用混合多克隆细胞亚群筛选和验证分析具有肺转移能力细胞亚群和基因。方法混合多克隆细胞亚群,分别尾静脉接种于喂饮强力霉素和对照两组裸小鼠,取小鼠肺原代培养得到具有肺转移能力的胰腺癌细胞亚群;提取基因组DNA进行PCR分析,比较两组小鼠特异基因检测率。结果体内筛选肺癌转移显示不加强力霉素组裸鼠出现肺转移症状早和严重。肺原代培养成功得到具有转移能力的胰腺癌细胞亚群,基因组DNA PCR分析表明75.0%没有喂饮强力霉素小鼠肺转移检测到SQSTM1基因,而在喂饮强力霉素小鼠只有37.5%肺转移检测到SQSTM1基因。PCR分析其他候选基因,包括sema4b,ube2k和gna13,均未检测到。证实强力霉素介导的SQSTM1基因表达导致胰腺癌细胞不同肺转移能力。结论多克隆混合同步筛选可验证得到胰腺癌肺转移细胞亚群和与其关联的基因。 展开更多
关键词 克隆同步筛选 胰腺癌 肿瘤转移 SQSTMI
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Characterization of the cDNA encoding a BPI/LBP homologue in venom gland of the hundred-pace snake Deinagkistrodon acutus 被引量:1
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作者 Jianrao HU Mingfu CAO Jiong CHEN 《Current Zoology》 SCIE CAS CSCD 北大核心 2009年第5期376-382,共7页
Bactericidal/permeability-increasing protein (BPI) and LPS-binding protein (LBP) play an important role in host defence. Current evidence shows that BPI/LBP may be widely existed in different cells and tissue type... Bactericidal/permeability-increasing protein (BPI) and LPS-binding protein (LBP) play an important role in host defence. Current evidence shows that BPI/LBP may be widely existed in different cells and tissue types of animals. A full-length cDNA clone encoding a BPI/LBP homologue (dBPI), 1757 bp in size, was characterized in venom gland of the hundred-pace snake Deinagkistrodon acutus. Its deduced amino acid sequence of 417 residues had 13.8% - 21.5% identity to BPI like 1 (BPIL1) and BPI like 3 (BPIL3) of other animals. Conserved cysteine residues which are involved in disulfide bond formation between the final strand of the N-terminal beta sheet and the long alpha helix of BPI are identified as Cys146-Cys183 of dBPI. Phylogenetic tree analysis showed that the BPI/LBP homologues formed five large clusters and dBPI was in a large cluster including BPIL1 and BPIL3. dBPI mRNA shows a tissue specific expression in venom gland. This is the first study to identify the cDNA encoding BPI/LBP homologues from reptiles [ Current Zoology 55 (5) : 376 - 382, 2009]. 展开更多
关键词 Hundred-pace snake Venom gland BPI/LBP homologue PHYLOGENY mRNA expression patterns
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Cloning of human brevican cDNA and expression of its mRNA in human glioma
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作者 韩晞 董艳 +2 位作者 由振东 何成 卢亦成 《Journal of Medical Colleges of PLA(China)》 CAS 2003年第6期388-392,共5页
Objective: To clone the cDNA of human brevican secreting isoform and to investigate its mRNA expression in human glioma. Methods: The full-length cDNA of human brevican secreted isoform was cloned from a human anaplas... Objective: To clone the cDNA of human brevican secreting isoform and to investigate its mRNA expression in human glioma. Methods: The full-length cDNA of human brevican secreted isoform was cloned from a human anaplastic astrocytoma by RT-PCR, and the expression of human brevican mRNA in 22 cases of human glioma and 13 cases of non-glial brain tumors were investigated by in situ hybridization. Results: The cDNA which including the whole open reading frame of human brevican secreted isoform was obtained. In situ hybridization showed that brevican positive cells were present in all of the 22 cases of gliomas (100%), whereas none were found in the 13 cases of non-glial and metastasis brain tumors examined. Conclusion: The results suggest that brevican mRNA is highly and specifically expressed in human glioma. 展开更多
关键词 BREVICAN RT-PCR in situ hybridization GLIOMA
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