Difference between revisions of "Part:BBa K3332027"

Revision as of 22:42, 27 October 2020

phnJ_mut45

The mutant of C-P lyase, which can degrade the glyphosate, from Enterobacterales. This subunit is essential to the activity of the whole enzyme.Use K823004 to construct a new part that can improve the capability of chassis bacteria to degrade glyphosate.

Biology

Phn system is a gene cluster for organophosphorus transport and degradation in many microorganisms. Enterobacterales use phnHIJK genes to encode C-P lyase, PhnJ protein is an essential subunit that can crack C-P bond. The 45th proline was mutated to glutamine.

Usage

We ligased the strong promoter、RBS、Terminator(BBa_J23100、BBa_B0034、BBa_B0015) and the parts (phnJ_mut45) on the expression vector pSB1C3 by standard assembly. Then the ligation mixture was transformed into E. coli DH5α & E. coli BL21(DE3), enabled the E. coli to degrade glyphosate at higher efficiency and improved the binding ability with the endogenous PhnHIK.

Characterization

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 549
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal NgoMIV site found at 597
Illegal AgeI site found at 268
Illegal AgeI site found at 775
1000
COMPATIBLE WITH RFC[1000]

Revision as of 17:52, 27 October 2020 (view source) Huangzinuo (Talk \| contribs) m ← Older edit		Revision as of 22:42, 27 October 2020 (view source) Master123 (Talk \| contribs) Newer edit →
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	===Usage===		===Usage===

−	We ligased the strong ~~promoter~~ (<partinfo>BBa_J23100</partinfo>) and the parts (~~RBS-~~phnJ_mut45~~-Terminator~~) on the expression vector pSB1C3 by standard assembly. Then the ligation mixture was transformed into ''E. coli'' DH5α & ''E. coli'' BL21(DE3), enabled the ''E. coli'' to degrade glyphosate at higher efficiency and improved the binding ability with the endogenous PhnHIK.	+	We ligased the strong promoter、RBS、Terminator(<partinfo>BBa_J23100</partinfo>、<partinfo>BBa_B0034</partinfo>、<partinfo>BBa_B0015</partinfo>) and the parts (phnJ_mut45) on the expression vector pSB1C3 by standard assembly. Then the ligation mixture was transformed into ''E. coli'' DH5α & ''E. coli'' BL21(DE3), enabled the ''E. coli'' to degrade glyphosate at higher efficiency and improved the binding ability with the endogenous PhnHIK.