Difference between revisions of "Part:BBa K3040005"

Revision as of 12:39, 20 October 2019 (view source) Liya (Talk | contribs) ← Older edit		Revision as of 12:46, 20 October 2019 (view source) Liya (Talk | contribs) Newer edit →
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	<partinfo>BBa_K3040005 short</partinfo>		<partinfo>BBa_K3040005 short</partinfo>
−	===Description===	+	==Description==
	It is pointed out by the previous iGEM team, UPF CRG Barcelona_2018 iGEM, that the native promoter pFadBA (BBa_K817002) has great leakage and is a weak promoter that isn’t ideal enough for our system. Therefore, to enhance the sensitivity and reduce the leakage of the promoter pFadBA for making it more suitable for our system, we modified the native promoter pFadBA by change consensus sequence of -10 and -35 region.		It is pointed out by the previous iGEM team, UPF CRG Barcelona_2018 iGEM, that the native promoter pFadBA (BBa_K817002) has great leakage and is a weak promoter that isn’t ideal enough for our system. Therefore, to enhance the sensitivity and reduce the leakage of the promoter pFadBA for making it more suitable for our system, we modified the native promoter pFadBA by change consensus sequence of -10 and -35 region.
	According to the previous research, there is improvement in expression for pFadBA with consensus sequence.		According to the previous research, there is improvement in expression for pFadBA with consensus sequence.
−	===Mechanism===	+	==Mechanism==
	Some study shows that in E. coli, there is sequence called consensus sequence which is universal and change the expression level of downstream protein. Consensus sequence is existing in -10 and -35 region, so it is relative with gene expression.		Some study shows that in E. coli, there is sequence called consensus sequence which is universal and change the expression level of downstream protein. Consensus sequence is existing in -10 and -35 region, so it is relative with gene expression.
	<br>		<br>
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	</html>		</html>
−	===Method===	+	==Method==
	1. Clone this promoter into DH5a		1. Clone this promoter into DH5a
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	<br>		<br>
−	===Result===	+	==Result==
	We detect the fluorescence by the mechanism. The following is the picture.		We detect the fluorescence by the mechanism. The following is the picture.
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	<!-- Add more about the biology of this part here-->		<!-- Add more about the biology of this part here-->
		+
		+	==Reference==
		+
		+	JENSEN, Peter Ruhdal; HAMMER, Karin. The sequence of spacers between the consensus sequences modulates the strength of prokaryotic promoters. Appl. Environ. Microbiol., 1998, 64.1: 82-87.
	===Usage and Biology===		===Usage and Biology===

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Revision as of 12:46, 20 October 2019

pFadBA promoter with consensus sequence regulating downstream RFP

Description

It is pointed out by the previous iGEM team, UPF CRG Barcelona_2018 iGEM, that the native promoter pFadBA (BBa_K817002) has great leakage and is a weak promoter that isn’t ideal enough for our system. Therefore, to enhance the sensitivity and reduce the leakage of the promoter pFadBA for making it more suitable for our system, we modified the native promoter pFadBA by change consensus sequence of -10 and -35 region. According to the previous research, there is improvement in expression for pFadBA with consensus sequence.

Mechanism

Some study shows that in E. coli, there is sequence called consensus sequence which is universal and change the expression level of downstream protein. Consensus sequence is existing in -10 and -35 region, so it is relative with gene expression.

The expression level can be change if sequence is modified. The patterns of the data points indicate which promoter clones contained mutant in either the -35 or the -10 region with consensus sequence.

Method

1. Clone this promoter into DH5a
2. Detection fluorescence level after adding different concentration of fatty acid.
3. Analysis the data

Result

We detect the fluorescence by the mechanism. The following is the picture.

The data below shows that pfadBA-NTHU promoter have 2.1 folds increase in expression over native pFadBA as the concentration of fatty acid rises after 4 hours. In conclusion, this promoter has higher fluorescence level in the higher fatty acid concentration environment.

This data shows the beginning fluorescence level of pFadBA_NTHU is much lower than pFadBA.

Actually, we improve the fold change of pFadBA promoter. Compare with the data from iGEM12_NTU-Taida, the promoter (BBa_K817002) only rise for about 1.5 fold change. However, after we modified the -10 and -35 region with consensus sequence, the fold change can be successfully increase.

Reference

JENSEN, Peter Ruhdal; HAMMER, Karin. The sequence of spacers between the consensus sequences modulates the strength of prokaryotic promoters. Appl. Environ. Microbiol., 1998, 64.1: 82-87.

Usage and Biology

Sequence and Features

Functional Parameters