Part:BBa_K1344004
malS-HlyA (with RBS)
This part is based on BBa_K523001, modified by deleting the stop codons. It's also fusion with HlyA secretional tag for extracelullar expression.
Starch Hydrolyzed Assay Using Iodine Test
Methods
Upon successful cloning of the three genes into our E.coli, we continued to confirm that all three genes are required have hydrolizing amylum activity on LB starch agar. We using iodine test to staining the amylum, and see the clearzone for some time variables.
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Discussion
We conducted starch hydrolyzed assay using three supernatant sample of E.coli on LB agar starch the flooding iodine on it.. We had two E. coli mutant, they are MH bacteria ( MalS and HlyA gene in pSB1C3 plasmid) and SMH bacteria (strong promoter constitutive-MalS-HlyA in pSB1C3), then we use E. coli Top10 wild type as negative control were growth on LB starch agar.
From the Assay result on 60 hours we can see if there were significantly different, SMH have the widest clear zone, next MH and E.coli wild type have the narrowest clear zone. SMH bacteria have the widest clear zone because the bacteria have an ability to secrete the enzyme into supernatant. But in MH bacteria they are enable to secrete enzyme and up take amylum as carbon source. On E.coli supernatant, the floating cell on supernatant aggregate become e new colony and growing on LB starch agar without have any clear zone or starch hydrolyzed activity
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1233
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 369
Illegal AgeI site found at 1534 - 1000COMPATIBLE WITH RFC[1000]
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