A combination method of the usual-PCR and reverse-PCR for the cloning of a novel lipase gene directly from the total genomic DNA of strain lip35 （Pseudomonas sp.） is described, whereby a lipase gene （lip） was clon...A combination method of the usual-PCR and reverse-PCR for the cloning of a novel lipase gene directly from the total genomic DNA of strain lip35 （Pseudomonas sp.） is described, whereby a lipase gene （lip） was cloned directly from genomic DNA. The sequence data have been deposited in the GenBank and EMBL data bank with the accession number EU414288. The nucleotide sequence showed a major open reading frame encoding a 59-kDa protein of 566 amino acid residues, which contained a lipase consensus sequence GXSXG. The lipase lip had 74 and 70% homologies with the lipases of an uncultured bacterium and P. fluorescens PfO-1, respectively, but it did not show any overall homology with lipases from other origins. The functional lipase was obtained when the lip gene was expressed in Pichia pastoris GS115.展开更多
[Objective] This study aimed to clone and analyze the sequence of CHS gene from Acer truncatum leaves. [Method] Using A. truncatum cultivars No.1-6 as experimental materials, total RNA was extracted from A. truncatum ...[Objective] This study aimed to clone and analyze the sequence of CHS gene from Acer truncatum leaves. [Method] Using A. truncatum cultivars No.1-6 as experimental materials, total RNA was extracted from A. truncatum leaves with the modified CTAB method. CHS gene sequences were downloaded from the NCBI and aligned by BLAST. Degenerate primers were designed by DNAMAN and Primer- premier5 to amplify the target band. CHS gene fragment was amplified by RT-PCR and ligated to pMD18-T vector. The identified positive colonies were sequenced. [Result] A 1 365 bp fragment was amplified. Sequence analysis suggested that the obtained fragment encoded 365 amino acids and shared above 90% homology to nucleotide sequence of CHS gene from A. palmatum and A. [Conclusion] In this study, CHS gene was successfully cloned from A. truncatum for the first time, which laid the foundation for efficient utilization of CHS gene.展开更多
To explore the structural characteristic of ribosomal protein S29 (rpS29) gene of Ailuropoda melanoleuca (giant panda), primers were designed based on the known nucleotide sequence of rpS29 genes to clone the cDNA and...To explore the structural characteristic of ribosomal protein S29 (rpS29) gene of Ailuropoda melanoleuca (giant panda), primers were designed based on the known nucleotide sequence of rpS29 genes to clone the cDNA and genomic sequences of this gene from giant panda by RT-PCR and PCR strategy respectively, and then the cloned cDNA and genomic sequences were sequenced and analyzed preliminarily. The results indicated that the cDNA fragment of the rpS29 from the giant panda is 205bp in size in length, containing an open reading frame (ORF) of 171bp, encoding 56 amino acids. The length of the genomic sequence is 1 598 bp, with three exons and two introns. The coding sequence shows a high degree of homology to those of Homo sapiens, Bos taurus, Rattus norvegicus and Mus musculus with 75%, 94.74%, 89.47% and 88.89% respectively, While all the homologies for amino acid sequences are high up to 100%. Primary structure analysis revealed that the molecular weight of the putative rpS29 protein is 6.68 KD, with a theoretical pI of 10.63. Based on topology prediction, there are three distinct types of functional sites in the rpS29 protein of giant panda.展开更多
基金The article was a part of the research program financed by the Science and Technology Bureau of Hebei Province, China (06220106D)
文摘A combination method of the usual-PCR and reverse-PCR for the cloning of a novel lipase gene directly from the total genomic DNA of strain lip35 （Pseudomonas sp.） is described, whereby a lipase gene （lip） was cloned directly from genomic DNA. The sequence data have been deposited in the GenBank and EMBL data bank with the accession number EU414288. The nucleotide sequence showed a major open reading frame encoding a 59-kDa protein of 566 amino acid residues, which contained a lipase consensus sequence GXSXG. The lipase lip had 74 and 70% homologies with the lipases of an uncultured bacterium and P. fluorescens PfO-1, respectively, but it did not show any overall homology with lipases from other origins. The functional lipase was obtained when the lip gene was expressed in Pichia pastoris GS115.
基金Supported by Agricultural Improved Variety Project of Shandong Province(LKZ[2014]No.96)
文摘[Objective] This study aimed to clone and analyze the sequence of CHS gene from Acer truncatum leaves. [Method] Using A. truncatum cultivars No.1-6 as experimental materials, total RNA was extracted from A. truncatum leaves with the modified CTAB method. CHS gene sequences were downloaded from the NCBI and aligned by BLAST. Degenerate primers were designed by DNAMAN and Primer- premier5 to amplify the target band. CHS gene fragment was amplified by RT-PCR and ligated to pMD18-T vector. The identified positive colonies were sequenced. [Result] A 1 365 bp fragment was amplified. Sequence analysis suggested that the obtained fragment encoded 365 amino acids and shared above 90% homology to nucleotide sequence of CHS gene from A. palmatum and A. [Conclusion] In this study, CHS gene was successfully cloned from A. truncatum for the first time, which laid the foundation for efficient utilization of CHS gene.
基金financially supported by Application Foundation Project in Sichuan Province(2009JY0061)Youth Fund Project of Educational Committee of Sichuan province(09ZB088)
文摘To explore the structural characteristic of ribosomal protein S29 (rpS29) gene of Ailuropoda melanoleuca (giant panda), primers were designed based on the known nucleotide sequence of rpS29 genes to clone the cDNA and genomic sequences of this gene from giant panda by RT-PCR and PCR strategy respectively, and then the cloned cDNA and genomic sequences were sequenced and analyzed preliminarily. The results indicated that the cDNA fragment of the rpS29 from the giant panda is 205bp in size in length, containing an open reading frame (ORF) of 171bp, encoding 56 amino acids. The length of the genomic sequence is 1 598 bp, with three exons and two introns. The coding sequence shows a high degree of homology to those of Homo sapiens, Bos taurus, Rattus norvegicus and Mus musculus with 75%, 94.74%, 89.47% and 88.89% respectively, While all the homologies for amino acid sequences are high up to 100%. Primary structure analysis revealed that the molecular weight of the putative rpS29 protein is 6.68 KD, with a theoretical pI of 10.63. Based on topology prediction, there are three distinct types of functional sites in the rpS29 protein of giant panda.
