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Team:Kyoto/Project/Goal C
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===Introduction=== | ===Introduction=== | ||
We checked the function of the anti-killer gene, SΔTMD1. | We checked the function of the anti-killer gene, SΔTMD1. | ||
| - | Lysis cassette encodes S gene and R gene and so | + | Killer gene, Lysis cassette encodes S gene and R gene and so and the transmembrane domein 1(TMD1) of this S gene is essential for the function of lysis cassette as killer-gene. |
| - | + | So SΔTMD1, which is a S mutant with TMD1 deleted, dominant-negatively inhibits Lysis cassette. | |
Then, after we made sure that this TMD1 is essential for lysis cassette, we checked if SΔTMD1 functions as anti-killer gene. | Then, after we made sure that this TMD1 is essential for lysis cassette, we checked if SΔTMD1 functions as anti-killer gene. | ||
| - | + | we used E.coli transformed with to check the importance of the TMD1 of S and E.coli transformed with to check the function of S&deltaTMD1. | |
These E.coli was grown in medium without IPTG and IPTG was added to the culture at proper time. The A550 of the culture was measured and the result was compared with that of the experiment of the killer gene in order to find whether the anti-killer gene works correctly. | These E.coli was grown in medium without IPTG and IPTG was added to the culture at proper time. The A550 of the culture was measured and the result was compared with that of the experiment of the killer gene in order to find whether the anti-killer gene works correctly. | ||
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| + | the lysis cassette allele with TMD1 of S gene deleted | ||
| + | lysis cassette regulated lac promoter and S&deltaTMD1 expressed constitutively | ||
===Method=== | ===Method=== | ||
Revision as of 17:36, 25 October 2010
Contents |
Goal C: Characterization of the anti-killer gene
Introduction
We checked the function of the anti-killer gene, SΔTMD1. Killer gene, Lysis cassette encodes S gene and R gene and so and the transmembrane domein 1(TMD1) of this S gene is essential for the function of lysis cassette as killer-gene. So SΔTMD1, which is a S mutant with TMD1 deleted, dominant-negatively inhibits Lysis cassette.
Then, after we made sure that this TMD1 is essential for lysis cassette, we checked if SΔTMD1 functions as anti-killer gene. we used E.coli transformed with to check the importance of the TMD1 of S and E.coli transformed with to check the function of S&deltaTMD1. These E.coli was grown in medium without IPTG and IPTG was added to the culture at proper time. The A550 of the culture was measured and the result was compared with that of the experiment of the killer gene in order to find whether the anti-killer gene works correctly.
the lysis cassette allele with TMD1 of S gene deleted
lysis cassette regulated lac promoter and S&deltaTMD1 expressed constitutively
Method
Bacterial strains
We used three types of E. coli, E. coli KRX transformed with <partinfo>BBa_K358021</partinfo>, KRX transformed with <partinfo>BBa_K358024</partinfo>, KRX transformed with <partinfo>BBa_K358022</partinfo>.
Measurement
We picked up three colonies from each plate, and cultivated them in M9 medium overnight. The overnight cultures were diluted to 0.1-0.12 (OD600) by M9 medium.After that, we added several amounts of IPTG to the cultures and measured OD600 of each culture at several points.
