human dentin to salivary phosphoprotein mature peptide cdna cloning sequence analysis and prokaryotic expression（人类牙质唾磷蛋白质成熟肽cdna克隆和原核表达序列分析）.docVIP

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2017-09-06 发布于浙江
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human dentin to salivary phosphoprotein mature peptide cdna cloning sequence analysis and prokaryotic expression（人类牙质唾磷蛋白质成熟肽cdna克隆和原核表达序列分析）.doc

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human dentin to salivary phosphoprotein mature peptide cdna cloning sequence analysis and prokaryotic expression（人类牙质唾磷蛋白质成熟肽cdna克隆和原核表达序列分析）

Human dentin to salivary phosphoprotein mature peptide cDNA cloning sequence Analysis and prokaryotic expression Author: Zhang Ying Hao Jianjun Shi Junnan, spoke Gu Shuping Wang Ping Jian Fei [Keywords] cloning Keywords: dentin protein; gene cloning; prokaryotic expression Abstract: Objective To clone human dentin phosphoprotein (DSPP mature peptide coding region of the gene fragments and prokaryotic expression. Guanidine isothiocyanate, one-step from human dental papilla tissue extraction of total RNA primer Oligo (dt) RT synthetic dental papilla cDNA, and then using the PCR method, amplified from the cDNA of the DSPP mature peptide gene fragment (approximately 600bp, The obtained gene fragment was inserted into the pBluescript plasmid vector, and transformed into E. coli XL1-Blue positive clones, extraction recombinant plasmid DNA, positive clones were identified by restriction analysis and nucleotide sequence analysis; DSPP gene was cloned into the expression vector pGEX-3X expression vector construction and expression in E. coli BL21. Results restriction map and sequence analysis consistent foreign literature, DSPP protein was expressed in E. coli. Conclusions DSPP mature peptide coding region of the gene fragment cloned into prokaryotic expression, and to lay the foundation for further study its function. Keywords: human dentin sailophosphoprotein; gene clone; prokaryotic expression Abstract: AIM To clone, sequence and prokaryotic express human dentin sailophosphoprotein (DSPP.METHODS In the study, total RNA was extracted from the tooth germs of a legally aborted embryo by acid guanidinium thiocyanata-phenol-choroform method, the desired DNA product was ob -tained from the total RNA by RT-PCR with the primers in-cluding Oligo (dtand two gene specific primers.The seg-ment (about600bpwas inserted into pBluescript vector and the interesting plasmid was transformed into E.coli host strain XL1-Blue. The double stranded DNA of the positive clone was a