Difference between revisions of "Part:BBa K1548000"

Latest revision as of 01:34, 18 October 2014

Hsp60 promoter (M. bovis)

Strong constitutive promoter from M. bovis which has also shown to function in a variety of mycobacteria. (1) It is expected to function in P. acnes a closely-related propionibacterium. The promoter does not function in E. coli, as we determined by an mRFP1 expression assay in the context of Part:BBa_K1548002, https://parts.igem.org/Part:BBa_K1548002 (Figure 1). This negative result was confirmed by E. coli promoter prediction software BPROM on (http://linux1.softberry.com/berry.phtml?topic=bprom&group=programs&subgroup=gfindb) which did not detect appropriate -10 and -35 E. coli promoter elements in the hsp60 promoter DNA sequence, validating our negative result.

Figure 1.The hsp60 promoter does not produce mRFP1 in E. coli. E. coli cells containing the hsp60-RBS-mRFP1 plasmid or hsp60-alone negative control plasmid were grown O/N and then diluted 1:250, grown for 4hrs diliuted to equal OD600 and assayed for mRFP1 fluoresecence in a fluorescence plate reader. n=3 and Stdev is plotted.

References: 1. Oldfield LM, Hatfull GF. Mutational Analysis of the Mycobacteriophage BPs Promoter PR Reveals Context-Dependent Sequences for Mycobacterial Gene Expression. J Bacteriol. 2014 Oct 15;196(20):3589-97. doi: 10.1128/JB.01801-14. Epub 2014 Aug 4.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 202
1000
COMPATIBLE WITH RFC[1000]

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 <partinfo>BBa_K1548000 short</partinfo>
-Strong constitutive promoter from ''M. bovis'' which has also shown to function in a variety of mycobacteria. (1) It is expected to function in ''P. acnes'' a closely-related propionibacterium. The promoter does not function in E. coli, as we determined by an mRFP1 expression assay. This result was confirmed by ''E. coli'' promoter prediction software BPROM on (http://linux1.softberry.com/berry.phtml?topic=bprom&group=programs&subgroup=gfindb) which did not detect appropriate -10 and -35 ''E. coli'' promoter elements in the hsp60 promoter DNA sequence, validating our negative result.
+Strong constitutive promoter from ''M. bovis'' which has also shown to function in a variety of mycobacteria. (1) It is expected to function in ''P. acnes'' a closely-related propionibacterium. The promoter does not function in E. coli, as we determined by an mRFP1 expression assay in the context of Part:BBa_K1548002, https://parts.igem.org/Part:BBa_K1548002 (Figure 1). This negative result was confirmed by ''E. coli'' promoter prediction software BPROM on (http://linux1.softberry.com/berry.phtml?topic=bprom&group=programs&subgroup=gfindb) which did not detect appropriate -10 and -35 ''E. coli'' promoter elements in the hsp60 promoter DNA sequence, validating our negative result.
+[[File:hsp60.png]]
+'''Figure 1.The hsp60 promoter does not produce mRFP1 in E. coli.''' ''E. coli'' cells containing the hsp60-RBS-mRFP1 plasmid or hsp60-alone negative control plasmid were grown O/N and then diluted 1:250, grown for 4hrs diliuted to equal OD600 and assayed for mRFP1 fluoresecence in a fluorescence plate reader. n=3 and Stdev is plotted.
 References:
 . Oldfield LM, Hatfull GF. Mutational Analysis of the Mycobacteriophage BPs Promoter PR Reveals Context-Dependent Sequences for Mycobacterial Gene Expression. J Bacteriol. 2014 Oct 15;196(20):3589-97. doi: 10.1128/JB.01801-14. Epub 2014 Aug 4.