Team:Warsaw/Calendar-Main/29 July 2008

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<img src="https://static.igem.org/mediawiki/2008/6/6a/Po_go.jpg" width=300/> <var><b>Fig. 1. Control SacI/NotI digests of isolated plasmids</b><br>  
<img src="https://static.igem.org/mediawiki/2008/6/6a/Po_go.jpg" width=300/> <var><b>Fig. 1. Control SacI/NotI digests of isolated plasmids</b><br>  
1. Marker<br>
1. Marker<br>
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2. digested pKS_omega_deltaA  
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2. digested pKS_omega_deltaA<br>
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3. digested pACYC_Omp_alpha
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3. digested pACYC_Omp_alpha<br></var>
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<br></var>
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Revision as of 23:20, 25 October 2008

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Cloning of omega_ΔA DNA fragment to pACYC177+OmpA_alpha

Michał K.

  1. Isolation of pKS+omega_ΔA from culture inoculated on previous day.
  2. Digest of pKS+omega_ΔA with SacI and NotI (BamHI buffer) (Fig.1.).
  3. Digest (SacI and NotI) and dephosphorylation (CIAP) of pACYC177+OmpA_alpha.
  4. Gel electrophoresis and gel-out of 4300 bp (pACYC177+OmpA_alpha) and 550 bp (omega_ΔA) bands.
  5. Ligation of DNA fragments from 2. and 3. (1 hr)
  6. Transformation of E. coli TOP10 strain with ligation.
  7. Transformants plating on LB + kanamycin.

Purification of proteins: Z-alpha and Z-omega

Piotr

pET15b+Z_alpha and pET15b+Z_omega in Rosetta strain inoculated in liquid LB with chloramphenicol and ampicillin.

Fig. 1. Control SacI/NotI digests of isolated plasmids
1. Marker
2. digested pKS_omega_deltaA
3. digested pACYC_Omp_alpha

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