Team:Warsaw/Calendar-Main/29 July 2008

From 2008.igem.org

(Difference between revisions)
 
(8 intermediate revisions not shown)
Line 5: Line 5:
<html>
<html>
 +
 +
 +
<h3>Purification of proteins: Z-alpha and Z-omega</h3><h4>Piotr</h4>
 +
 +
<p><A href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BZ-alpha>pET15b+Z_alpha</A> and <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2Bhis%2BZ%2Bomega>pET15b+Z_omega</a> in <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#rosetta">Rosetta</a> strain inoculated in liquid LB with chloramphenicol and ampicillin.
 +
</p>
<h3>Cloning of omega_&Delta;A DNA fragment to <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-alpha>pACYC177+OmpA_alpha</a></h3><h4>Michał K.</h4>
<h3>Cloning of omega_&Delta;A DNA fragment to <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-alpha>pACYC177+OmpA_alpha</a></h3><h4>Michał K.</h4>
<ol>
<ol>
<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pKSII%2Bomega-deltaA>pKS+omega_&Delta;A</a> from culture inoculated on previous day.</li>
<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pKSII%2Bomega-deltaA>pKS+omega_&Delta;A</a> from culture inoculated on previous day.</li>
-
<li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest>Digest</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pKSII%2Bomega-deltaA>pKS+omega_&Delta;A</a>  with SacI and NotI (BamHI buffer) (Fig.1.).</li>
+
<li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest>Digest</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pKSII%2Bomega-deltaA>pKS+omega_&Delta;A</a>  with SacI and NotI (BamHI buffer).</li>
<li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest>Digest</a> (SacI and NotI) and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#removing">dephosphorylation</a> (CIAP) of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-alpha>pACYC177+OmpA_alpha</a>. </li>
<li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest>Digest</a> (SacI and NotI) and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#removing">dephosphorylation</a> (CIAP) of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-alpha>pACYC177+OmpA_alpha</a>. </li>
-
<li>Gel electrophoresis and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of 4300 bp (<a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-alpha>pACYC177+OmpA_alpha</a>) and 550 bp (omega_&Delta;A) bands. </li>
+
<li>Gel electrophoresis and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of 4300 bp (<a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-alpha>pACYC177+OmpA_alpha</a>) and 550 bp (omega_&Delta;A) bands (<a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/29_July_2008#fig1">Fig. 1.</a>).</li>
-
<li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">Ligation</a> of DNA fragments from 2. and 3. (1 hr)</li>
+
<li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">Ligation</a> of DNA fragments from 2. and 3. (1 hr).</li>
<li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#chemotransform">Transformation</a> of E. coli <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">TOP10</a> strain with ligation.</li>
<li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#chemotransform">Transformation</a> of E. coli <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">TOP10</a> strain with ligation.</li>
<li>Transformants plating on LB + kanamycin.</li></ol>  
<li>Transformants plating on LB + kanamycin.</li></ol>  
</p>
</p>
-
 
+
<a name="fig1"><img src="https://static.igem.org/mediawiki/2008/6/6a/Po_go.jpg" width=200/></a> <var><b>Fig. 1. </b> SacI/NotI digests of isolated plasmids<br>  
-
<h3>Purification of proteins: Z-alpha and Z-omega</h3><h4>Piotr</h4>
+
-
 
+
-
<p><A href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BZ-alpha>pET15b+Z_alpha</A> and <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2Bhis%2BZ%2Bomega>pET15b+Z_omega</a> in <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#rosetta">Rosetta</a> strain inoculated in liquid LB with chloramphenicol and ampicillin.
+
-
</p>
+
-
 
+
-
<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>
-
2. digested pKS_omega_deltaA
+
2. digested pKS_omega_&Delta;A<br>
-
3. digested pACYC_Omp_alpha
+
3. digested pACYC_Omp_alpha<br></var>
-
<br></var>
+

Latest revision as of 16:55, 28 October 2008

Gallery Bricks Notebook Team Project Home


Previous day
return to main notebook page
Previous entry
Interactive list of topics
next notebook entry

 



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.

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).
  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 (Fig. 1.).
  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.

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

Retrieved from "http://2008.igem.org/Team:Warsaw/Calendar-Main/29_July_2008"