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- W2367654205 abstract "Objective It is to construct the recombinant plasmid vector that codes calcitonin gene-related peptide(CGRP) of rat(pcDNA3.1(+)/rCGRP). Methods The total RNA was extracted from the brain tissue of rat with Trizol reagent,and the cDNA of rCGRP was amplified by RT-PCR(PCR primers were designed with double restriction enzyme sites).The PCR products and pcDNA3.1(+) were digested with restriction enzymes,purified and ligated.Recombinant colonies were harvested and identified by PCR,restriction enzyme digestion and DNA sequencing. Results(1)Electrophoretic analysis of RT-PCR product: under the 1% agarose gel electrophoresis,the amplified products of RT-PCR presented a specific band that was coincided with the size of rCGRP cDNA;(2)Identification of the recombinant colony: ①Identification with PCR Under the agarose gel electrophoresis,the amplified products of PCR presented a specific band that coincided with the size of rCGRP cDNA;②Identification with restricted digestion.The extracted plasmids were digested by two restriction enzymes,and the digested products presented two specific bands those coincided with the sizes of pcDNA3.1 and rCGRP cDNA respectively;③DNA sequencing The recombinant plasmid identified by the former steps was sent out for direct DNA sequencing.The acquired sequence was analyzed by BLAST and demonstrated to be completely identical with the coding region sequence of rCGRP. Conclusion(1)There exists gene expression of CGRP in the brain tissue of rat;(2)The pcDNA3.1(+)/rCGRP is successfully constructed with the techniques of RT-PCR and gene recombination." @default.
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- W2367654205 date "2008-01-01" @default.
- W2367654205 modified "2023-09-28" @default.
- W2367654205 title "Construction of eukaryotic expression vector coding rat calcitonin gene-related peptide(pcDNA3.1/rCGRP)" @default.
- W2367654205 hasPublicationYear "2008" @default.
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