Difference between revisions of "Part:BBa K4137009"
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<partinfo>BBa_K4137009 short</partinfo> | <partinfo>BBa_K4137009 short</partinfo> | ||
− | + | This part synthesizes and secretes a novel bacterial chitinase Chi18H8, which can be released extracellularly through e.coli Sec pathway (SecB-SecA-SecYEG, OmpF). This system continuously promotes the overexpression of Chi18H8 to lyse the chitin cell wall of the fungus fusarium oxysporum f.sp cubense, inhibiting fungal proliferation. | |
− | + | [[File:t7-yebf-gs-chitinase.png|800px|thumb|center|Fig.1 Chitinase (chi18h8) SEC secretion construct.]] | |
− | === | + | |
+ | ===Construct Designs=== | ||
+ | |||
+ | We attached a 6x His-Tag downstream of the chi18h8 sequence for purification purposes. Attached to the upstream of the chi18h8 sequence is a glycine-serine linker (GS linker) (BBa_J18921) that links chi18h8 to a yebF secretion tag (BBa_K2922002), which forms our open reading frame (ORF). We used a T7 strong promoter and flanked RBS (AAGGAG) upstream of the ORF. The terminator BBa_B1006 is attached downstream of the ORF. | ||
+ | |||
+ | ===Characterization=== | ||
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Revision as of 05:49, 9 October 2022
T7+RBS+YebF-GS Linker-chi18h8-6xHis+B1006
This part synthesizes and secretes a novel bacterial chitinase Chi18H8, which can be released extracellularly through e.coli Sec pathway (SecB-SecA-SecYEG, OmpF). This system continuously promotes the overexpression of Chi18H8 to lyse the chitin cell wall of the fungus fusarium oxysporum f.sp cubense, inhibiting fungal proliferation.
Construct Designs
We attached a 6x His-Tag downstream of the chi18h8 sequence for purification purposes. Attached to the upstream of the chi18h8 sequence is a glycine-serine linker (GS linker) (BBa_J18921) that links chi18h8 to a yebF secretion tag (BBa_K2922002), which forms our open reading frame (ORF). We used a T7 strong promoter and flanked RBS (AAGGAG) upstream of the ORF. The terminator BBa_B1006 is attached downstream of the ORF.
Characterization
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1523
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 1036
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 583
- 1000COMPATIBLE WITH RFC[1000]