Team:Kyoto/Project/Goal C

Goal C: Characterization of the anti-killer gene

Introduction

We checked the function of the anti-killer gene, SΔTMD1. Lysis cassette encodes S gene, R gene and so on 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.(check Learn more)

Then, we made sure that this TMD1 is essential for lysis cassette and checked if SΔTMD1 functions as anti-killer gene. To check the importance of the TMD1, we checked if a lysis cassette allele with TMD1 of S gene deleted (lysis cassetteΔTMD1) functions as killer-gene. To research the function of SΔTMD1, we checked if SΔTMD1 can prevent lysis cassette from working as killer-gene in E.coli transformed with both these genes. Because E.coli is immediately dead if killer-gene is constitutively expressed, we made lysis cassette regulated by lac promoter, after E.coli was grown lysis cassette is expressed by IPTG and we checked if E.coli were alive due to anti-killer gene, SΔTMD1 expressed constitutively .

Experiment 1 Function check of SΔTMD1

Bacterial strains

We used two types of E. coli, E. coli KRX transformed with <partinfo>BBa_K358019</partinfo>, KRX transformed with <partinfo>BBa_K358022</partinfo>.

Measurement

We picked out three colonies from each devices, and cultivated them in M9 medium with 1mM IPTG and without IPTG overnight. Then, we measured A550 of each cultures.

Result

Result

Discussion