TAILIEUCHUNG - pXST, a novel vector for TA cloning and blunt-end cloning

With the rapid development of sequencing technologies, increasing amount of genomic information has been accumulated. To clone genes for further functional studies in large scale, a cheap, fast and efficient cloning vector is desired. | Liu et al. BMC Biotechnology 2018 18 44 https s12896-018-0456-8 BMC Biotechnology METHODOLOGY ARTICLE Open Access pXST a novel vector for TA cloning and blunt-end cloning 1 1 11 112 1 Qin Liu Hui-Jie Dang Yuan-Hang wu Min LI Yin-Hua Chen Xiao-Lei Niu Kai-Mian LI ano Li-Juan Luo Abstract Background With the rapid development of sequencing technologies increasing amount of genomic information has been accumulated. To clone genes for further functional studies in large scale a cheap fast and efficient cloning vector is desired. Results A bifunctional vector pXST has been constructed. The pXST vector harbors a XcmI-ccdB-XcmI cassette and restriction site Smal. Digestion the vector with Xcml generates a single thymidine T overhang at 3 end which facilitates TA cloning and Smal gives blunt end that enables the blunt-end ligation. Multiple products with various sizes were amplified from cassava genome by PCR and each PCR fragment was separately cloned into a pXST using TA cloning and blunt-end ligation methods. In general the TA cloning gave higher transformation efficiency than blunt-end ligation for inserts with all different sizes and the transformation efficiency significantly decreased with increasing size of inserts. The highest transformation efficiency X 106 transformants pg was achieved when cloning 517 bp DNA fragment using TA cloning. No significant difference observed in the positive cloning efficiency between two ligation methods and the positive cloning efficiency could reach as high as 100 especially for small inserts . 517 and 957 base pairs . Conclusions We describe a simple and general method to construct a novel pXST vector. We confirm the feasibility of using pXST vector to clone PCR products amplified from cassava genome with both TA cloning and blunt-end ligation methods. The pXST plasmid has several advantages over many currently available vectors in that 1 it possesses Xcml-ccdB-Xcml cassette and restriction site Smal .

• Sinh học
• BMC Biotechnology
• PCR product
• T vector
• Blunt end ligation
• High efficiency
• Transgenic animals
• Recombinant proteins
• Animal biotechnology
• Functional cytokines
• ABC transporter
• Taxonomic diversity
• ATP binding cassette
• Transporter gene superfamily
• Community engagement
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• Global health
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• BMC Chemistry
• Citric acid
• White biotechnology
• Organic acid
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• MicroRNAs
• cDNA synthesis
• synthetic templates
• Design of PCR primers
• Quantitative PCR
• Error prone PCR
• Site directed mutation
• Site saturation mutagenesis
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• Tobacco stalk
• Nicotine degradation
• Phanerochaete chrysosporium
• Lignocellulolytic enzymes
• Chaetomium globosum
• Synergistic action
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• Phenolic alcohols
• β xylosidase
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• Geobacillus sp
• Hydrolyzing xylo oligosaccharides
• Heterologous expression
• Kluyveromyces lactis
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