Difference between revisions of "Part:BBa K2243033"
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To test the influence of attBP sites of TP901-1 to terminator ECK120030221 (abbreviation: 221) in the forward direction. | To test the influence of attBP sites of TP901-1 to terminator ECK120030221 (abbreviation: 221) in the forward direction. | ||
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| − | Usage | + | <h2>Usage</h2> |
We constructed this part to characterize the recombination efficiency of the recombinase Lactococcal phage TP901-1. It consists of the terminator ECK120030221 (abbreviation: 221) in the forward direction flanked by attB and attP sites of recombinase TP901-1. Upon recombination, the orientation of the terminator changes. As a result, expression of downstream sequence is blocked. | We constructed this part to characterize the recombination efficiency of the recombinase Lactococcal phage TP901-1. It consists of the terminator ECK120030221 (abbreviation: 221) in the forward direction flanked by attB and attP sites of recombinase TP901-1. Upon recombination, the orientation of the terminator changes. As a result, expression of downstream sequence is blocked. | ||
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| − | Biology | + | <h2>Biology</h2> |
The attP site of TP901-1 is used to integrate phage DNA at the host attB site of Lactococcal 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. | The attP site of TP901-1 is used to integrate phage DNA at the host attB site of Lactococcal 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. | ||
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We first characterized the terminator strength using the following formula: | We first characterized the terminator strength using the following formula: | ||
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[[File:Peking TS good terminators.png|600px|thumb|center|Forward and Reverse Terminator Strength Ratio Induced with 0.1mM and 1mM IPTG]] | [[File:Peking TS good terminators.png|600px|thumb|center|Forward and Reverse Terminator Strength Ratio Induced with 0.1mM and 1mM IPTG]] | ||
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| − | Terminator Reference Table | + | <h2>Terminator Reference Table</h2> |
[[Media:Peking_trt.xlsx]] | [[Media:Peking_trt.xlsx]] | ||
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Revision as of 15:35, 1 November 2017
TP901-1 attB_221F_TP901-1 attP
To test the influence of attBP sites of TP901-1 to terminator ECK120030221 (abbreviation: 221) in the forward direction.
Usage
We constructed this part to characterize the recombination efficiency of the recombinase Lactococcal phage TP901-1. It consists of the terminator ECK120030221 (abbreviation: 221) in the forward direction flanked by attB and attP sites of recombinase TP901-1. Upon recombination, the orientation of the terminator changes. As a result, expression of downstream sequence is blocked.
Biology
The attP site of TP901-1 is used to integrate phage DNA at the host attB site of Lactococcal 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]