Team:Newcastle/9 July 2010

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===Transformation===
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{{Team:Newcastle/mainbanner}}
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Link to transformation protocol (to be added)
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=Research=
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Today we initiated with our research in urease which would lead to calcium carbonate production inside the crack.
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====Tubes used:====
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#Wray LV, Ferson E and Fisher SH. (1997). "Expression of the Bacillus subtilis ureABC operon is controlled by multiple regulatory factors including CodY, GlnR, TnrA, and Spo0H". ''Journal of bacteriology''. 179(17). 5494-501.
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Each of the following five tubes contain 200 µl of competent ''E. coli'' DH5alpha. To this the DNA to be transformed was added.
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#Kim JK, Mulrooney SB and Hausinger RP. (2005). "Biosynthesis of Active Bacillus subtilis Urease in the Absence of Known Urease Accessory Proteins". ''Journal of Bacteriology''. 187(20). 7150-7154.
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# 1:3 (contains LacI insert and pSB1AT3 vector in the proportion of 1:3)
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# 1:5 (contains LacI insert and pSB1AT3 vector in the proportion of 1:5)
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# Negative control for ligation (contains vector with no insert)
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# Control for transformation (without plasmid)
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# Control for transformation (with plasmid, pSB1AT3)
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====Protocol:====
 
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# Thaw a 200 µl aliquot of ''E. coli'' DH5alpha and add the transforming DNA. For tubes 1-3 5 µl of vector was added, for tube 4 no vector was added and for tube 5 only 1 µl of vector was added. This is because tubes 1 to 3 have been ligated and will contain cells in different forms of the ligated vector whereas tube 5 contains the ligated vector (our control).
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{{Team:Newcastle/footer}}
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# Incubate for 45 mins on ice.
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# Heat-shock the cells at 42°C for 120 secs, and place on ice again for 3-4 min.
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# Add 1 ml of LB broth to each tube and incubate the cells at 37°C for 1.5 hr (static for initial 20 mins, shaking for remaining time).
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# Plate out 200 µl/plate on LB (agar at 1.5%).
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#*
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# Incubate plates overnight at 37°C.
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Latest revision as of 19:16, 25 October 2010

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Research

Today we initiated with our research in urease which would lead to calcium carbonate production inside the crack.

  1. Wray LV, Ferson E and Fisher SH. (1997). "Expression of the Bacillus subtilis ureABC operon is controlled by multiple regulatory factors including CodY, GlnR, TnrA, and Spo0H". Journal of bacteriology. 179(17). 5494-501.
  2. Kim JK, Mulrooney SB and Hausinger RP. (2005). "Biosynthesis of Active Bacillus subtilis Urease in the Absence of Known Urease Accessory Proteins". Journal of Bacteriology. 187(20). 7150-7154.


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