Difference between revisions of "Part:BBa K4907112"
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===Biology=== | ===Biology=== | ||
====pVSW-3(GGG)==== | ====pVSW-3(GGG)==== | ||
− | Some RNA polymerases of eukaryotes and viruses have domains that specifically recognize DNA base sequences, and they are specifically matched with their corresponding promoters(1). VSW-3 RNAP is encoded by the chillophilic phage VSW-3 in plateau lakes and has low temperature specificity(2). Hengxia <i>et al</i>. characterized pVSW-3 series promoters for the first time and pVSW-3(GGG) is one of them. | + | Some RNA polymerases of eukaryotes and viruses have domains that specifically recognize DNA base sequences, and they are specifically matched with their corresponding promoters (1). VSW-3 RNAP is encoded by the chillophilic phage VSW-3 in plateau lakes and has low-temperature specificity (2). Hengxia <i>et al</i>. characterized pVSW-3 series promoters for the first time and pVSW-3(GGG) is one of them. |
===Usage and design=== | ===Usage and design=== | ||
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1.S. Borukhov, E. Nudler, RNA polymerase: the vehicle of transcription. <i>Trends in Microbiology</i> <b>16</b>, 126-134 (2008). | 1.S. Borukhov, E. Nudler, RNA polymerase: the vehicle of transcription. <i>Trends in Microbiology</i> <b>16</b>, 126-134 (2008). | ||
− | 2. H. Xia <i>et al.</i>, Psychrophilic phage VSW-3 RNA polymerase reduces both terminal and full-length dsRNA byproducts in in vitro transcription. <i>RNA Biology</i> <b>19</b>, 1130-1142 (2022). | + | 2. H. Xia <i>et al.</i>, Psychrophilic phage VSW-3 RNA polymerase reduces both terminal and full-length dsRNA byproducts in <i>in vitro</i> transcription. <i>RNA Biology</i> <b>19</b>, 1130-1142 (2022). |
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Latest revision as of 10:12, 11 October 2023
pVSW-3(GGG)-B0034-gfp-B0015
Biology
pVSW-3(GGG)
Some RNA polymerases of eukaryotes and viruses have domains that specifically recognize DNA base sequences, and they are specifically matched with their corresponding promoters (1). VSW-3 RNAP is encoded by the chillophilic phage VSW-3 in plateau lakes and has low-temperature specificity (2). Hengxia et al. characterized pVSW-3 series promoters for the first time and pVSW-3(GGG) is one of them.
Usage and design
XMU-China has developed a novel RNA polymerase, VSW-3 RNAP and we characterized its potentially useful promoters in order to construct a matching expression system. pVSW-3(GGG) is one of the series promoters. BBa_K4907112_pSB3K3 was constructed as a reporting circuit, for comparing with pVSW-3(18) and pVSW-3(genome). By characterizing these three promoters, we hope to determine the effect of the 3' terminal structure of the promoter for VSW-3 RNAP on its efficiency, and to identify a VSW-3 expression system that can effectively function in E. coli.
![](https://static.igem.wiki/teams/4907/wiki/parts/jincheng/biaozhen/pvsw-3-all-gfp.png)
Characterization
Agarose gel electrophoresis (AGE)
When we were building this circuit, colony PCR was used to certify the plasmid was correct. We got the target fragment-1220bp (lane K4907112).
![](https://static.igem.wiki/teams/4907/wiki/parts/jincheng/biaozhen/112-1.png)
Comparison of series promoters: pVSW-3(GGG), pVSW-3(genome)
In order to find a promoter that can function efficiently in Escherichia coli, we constructed BBa_K4907109_pSB3K3(pVSW-3(18)), BBa_K4907112_pSB3K3(pVSW-3(GGG)) and BBa_K4907122_pSB3K3(pVSW-3(genome)) to explore the effect of the structure of the 3' terminal of the promoter on its efficiency. The results are shown in the figure, with BBa_K4907109_pSB3K3 showing the highest efficiency.
![](https://static.igem.wiki/teams/4907/wiki/parts/jincheng/biaozhen/ggg-gemone.png)
Reference
1.S. Borukhov, E. Nudler, RNA polymerase: the vehicle of transcription. Trends in Microbiology 16, 126-134 (2008).
2. H. Xia et al., Psychrophilic phage VSW-3 RNA polymerase reduces both terminal and full-length dsRNA byproducts in in vitro transcription. RNA Biology 19, 1130-1142 (2022).
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]