31228020)Ĭompeting interests: The authors have declared that no competing interests exist. 58-4) and National Natural Science Foundation of China (contract no. The work is made available under the Creative Commons CC0 public domain dedicationĭata Availability: All relevant data are within the paper and its Supporting Information files.įunding: This research is supported by the US Department of Agriculture (US–Egypt Science and Technology Joint Fund project no.
#Sv40 poly a snapgene free#
This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. Received: SeptemAccepted: NovemPublished: December 14, 2015
PLoS ONE 10(12):Įditor: Yi Li, Wuhan Bioengineering Institute, CHINA This work suggests that SV40 polyA in BEVSs should be replaced by an AcMNPV late gene polyA for optimal protein production or left untouched for optimal RNA production (RNA interference applications).Ĭitation: Salem TZ, Seaborn CP, Turney CM, Xue J, Shang H, Cheng X-W (2015) The Influence of SV40 polyA on Gene Expression of Baculovirus Expression Vector Systems. Therefore, we conclude that SV40 polyA increases mRNA levels but decreases protein production in the BEVS when the polyhedrin promoter is used at different loci. In this study, spectrofluorometry and western blot showed reduction of EGFP protein for all recombinant viruses with SV40 polyA, whereas qPCR showed an increase in the egfp mRNA levels. In order to test the significance of adding the SV40 polyA sequence on gene expression, the expression of the enhanced green fluorescent protein (egfp) was evaluated with and without the presence of SV40 polyA under the control of the polyhedrin promoter at different genomic loci (polyherin, ecdysteroid UDP-glucosyltransferase (egt), and gp37). Moreover, the effect of the SV40 polyA sequence on the polyhedrin promoter activity has not been tested either at its natural polyhedrin locus or in other loci in the viral genome. In the baculovirus prototype Autographa californica multiple nucleopolyhedrovirus (AcMNPV), the polyhedrin promoter (very late promoter) transcribes its gene by a viral RNA polymerase therefore there is no supporting evidence that SV40 polyA is required for the proper gene expression under the polyhedrin promoter. Codons of the type NUC, NCG and CGN are absent or very rare.The simian virus 40 polyadenylation signal (SV40 polyA) has been routinely inserted downstream of the polyhedrin promoter in many baculovirus expression vector systems (BEVS). The use of degenerate codons is decidedly non-random, but is similar for the early and late regions. The mRNAs for the latter three proteins are presumably spliced out of a common primary RNA transcript. The almost complete amino acid sequences of the two early proteins as well as those of the late proteins have been deduced from the nucleotide sequence. In the late region the gene for the major protein VP1 overlaps those for proteins VP2 and VP3 over 122 nucleotides but is read in a different frame. Particular points of interest revealed by the complete sequence are the initiation of the early t and T antigens at the same position and the fact that the T antigen is coded by two non-contiguous regions of the genome the T antigen mRNA is spliced in the coding region. At least 15.2% of the genome is presumably not translated into polypeptides. The determination of the total 5,224 base-pair DNA sequence of the virus SV40 has enabled us to locate precisely the known genes on the genome.
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