Difference between revisions of "Part:BBa K2243032"
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===Characterization=== | ===Characterization=== | ||
− | + | We first characterized the terminator strength using the following formula: | |
Ts=〖GFP〗of the random sequence/〖[GFP]〗with the terminator | Ts=〖GFP〗of the random sequence/〖[GFP]〗with the terminator | ||
+ | [[File:Peking_TS_0.1mM.png|800px|thumb|center|Terminator Strength Induced with 0.1mM IPTG]] | ||
+ | |||
+ | [[File:Peking_TS_1mM.png|800px|thumb|center|Terminator Strength Induced with 1mM IPTG]] | ||
And we sifted out 6 desirable unidirectional terminators without potential cryptic promotor in both orientations. | And we sifted out 6 desirable unidirectional terminators without potential cryptic promotor in both orientations. | ||
− | [[File:Peking | + | [[File:Peking ratio.png|600px|thumb|center|Terminator Strength Induced with 0.1M IPTG]] |
+ | |||
+ | [[File:Peking TS good terminators.png|600px|thumb|center|Forward and Reverse Terminator Strength Ratio Induced with 0.1mM and 1mM IPTG]] | ||
+ | |||
− | |||
− | + | <!-- --> | |
+ | Terminator Reference Table | ||
− | + | [[Media:Peking_trt.xlsx]] | |
− | |||
Revision as of 14:44, 1 November 2017
phiC31 attB_435R_phiC31 attP
To test the influence of attBP sites of phiC31 to terminator ECK120034435 (abbreviation: 435) in the reverse direction.
Usage and Biology
Usage
We constructed this part to characterize the recombination efficiency of the recombinase Streptomyces bacteria phage phiC31. It consists of the terminator ECK120034435 (abbreviation: 435) in the reverse direction flanked by attB and attP sites of recombinase phiC31. Upon recombination, the orientation of the terminator changes. As a result, expression of downstream sequence is blocked.
Biology
The attP site of phiC31 is used to integrate phage DNA at the host attB site of Streptomyces bacterium smegmatis, generating the recombinant junctions attL and attR. DNA cleavage and re-ligation occur at the central crossover region at attB and attP, which allows the sequence to be flipped, excised, or inserted between recognition sites. We obtained the terminator, attB and attP sites by oligo synthesis.
Characterization
We first characterized the terminator strength using the following formula:
Ts=〖GFP〗of the random sequence/〖[GFP]〗with the terminator
And we sifted out 6 desirable unidirectional terminators without potential cryptic promotor in both orientations.
Terminator Reference Table
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]