Difference between revisions of "Part:BBa K3924047"
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==Design and Construction== | ==Design and Construction== | ||
To construct the plasmid, we use endonuclease SpeI to cut RGP-mchE at 2bp after mchE open reading frame(ORF), then we used NEB Hifi DNA assembly to add RBS(ribosome binding site)-mchF between mchE and rrnB terminator.<br/> | To construct the plasmid, we use endonuclease SpeI to cut RGP-mchE at 2bp after mchE open reading frame(ORF), then we used NEB Hifi DNA assembly to add RBS(ribosome binding site)-mchF between mchE and rrnB terminator.<br/> | ||
| − | [[Image: T--Tsinghua--part_mchEF_construct.png|'''Figure 1: The design of mchEF''']] | + | [[Image: T--Tsinghua--part_mchEF_construct.png|center|600px|thumb|'''Figure 1: The design of mchEF''']] |
Revision as of 09:57, 21 October 2021
mchE-mchF polycistrons with Ptac lacO promoter
mchE-mchF polycistrons with Ptac lacO promoter
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 1713
Illegal PstI site found at 610
Illegal PstI site found at 823 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 1713
Illegal PstI site found at 610
Illegal PstI site found at 823 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 1713
Illegal BamHI site found at 237
Illegal BamHI site found at 2380 - 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 1713
Illegal PstI site found at 610
Illegal PstI site found at 823 - 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 1713
Illegal PstI site found at 610
Illegal PstI site found at 823
Illegal AgeI site found at 2761 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 1347
Illegal SapI site found at 2456
Profile
Name: mchEF
Base Pairs: 3298bp
Origin: Escherichia coli, merge mchE and mchF
Properties: A polycistron can express both MchE and MchF. MchE and MchL transport mature MccH47 to the extracellular side.
Usage and Biology
The mchE and mchF are both from E.coli CA46. The mchE encodes MchE and the mchF encodes MchF. Both MchE and MchF transport mature MccH47 to the extracellular side.[1] The polycistron aim to make the engineering bacteria have the ability to transport MccH47.
Design and Construction
To construct the plasmid, we use endonuclease SpeI to cut RGP-mchE at 2bp after mchE open reading frame(ORF), then we used NEB Hifi DNA assembly to add RBS(ribosome binding site)-mchF between mchE and rrnB terminator.
Reference
[1] Vassiliadis G, Destoumieux-Garzón D, Lombard C, Rebuffat S, Peduzzi J. Isolation and characterization of two members of the siderophore-microcin family, microcins M and H47. Antimicrobial Agents and Chemotherapy. 2010 Jan;54(1):288-297. DOI: 10.1128/aac.00744-09. PMID: 19884380; PMCID: PMC2798501.