July

From 2008.igem.org

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Revision as of 19:39, 9 October 2008

07-04-2008

Assess Pool
We mixed all oligos together, except for the modified, the left and the right borders and the remainders:

Oligos modified with flourophor: 1. board:
r-3t8f
r-5t10e
r-5t12e

Oligos modified with NIP: 1. board:
r-7t6f
r-7t8f
r-7t10f
r-5t8f
r-3t10e
r-7t6e
3. board:
r7t22e

Left border: 1.board, 1.line
  Right border: 3.board, 2.line
  Remainders: 3.board, 3.line

07-06-2008
Assessment of cell-culture
-(DYT-Medium; ER 2738-cells)
50ml DYT-Medium
+50 TET (25ng/ml -> 1:1000)
+ pick ER 2738-cells (from -80 degree freezer)
-> shake at 37 degree over night

07-07-2008
1) Thin down overnight culture to OD~0.1
1:10 -> 0.66 = OD 660

5000µl = 0.66
x = 0.1 -> x = 758µl ~ 760µl in 50ml
 We mixed 760µl cell culture with 50ml DYT and shaked both at 37°C; Inoculation after 95min
<p>2) Inoculation of cell culture with M13mp18 phages
~ 50ml cell culture
+ 5µl M13mp18 phages (from -80°C)
-> 4h shaking at 37°C

3) Phage precipitation
1) centrifuging of the phage-solution at 5000 x g for 20 min.
2) after decanting supernatant dessolve the pellet in 7ml (1/7 of the supernatant volume) PEG/NaCl
-> The precipitation occurs at 4°C over night

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 {{Freiburg2008_Main|
 Content=
-under construction
+<p>'''07-04-2008'''<br />
+  <br />
+  '''Assess Pool'''<br />
+  We mixed all oligos together, except for the modified, the left and the right borders and the remainders:<br />
+  <br />
+  '''Oligos modified with flourophor:'''                     1. board: <br />
+  r-3t8f<br />
+  r-5t10e<br />
+  r-5t12e<br />
+  <br />
+  '''Oligos modified with NIP:'''                              1. board:<br />
+  r-7t6f<br />
+  r-7t8f<br />
+  r-7t10f<br />
+  r-5t8f<br />
+  r-3t10e<br />
+  r-7t6e<br />
+. board:<br />
+  r7t22e<br />
+  <br />
+  '''Left border:''' 1.board, 1.line<br />   '''Right border:''' 3.board, 2.line<br />   '''Remainders:''' 3.board, 3.line<br />
+  <br />
+  '''07-06-2008'''<br />
+  '''Assessment of cell-culture'''<br />
+  -(DYT-Medium; ER 2738-cells)<br />
+ml            DYT-Medium<br />
+  +50             TET (25ng/ml -&gt; 1:1000)<br />
+  +                 pick ER 2738-cells (from -80 degree freezer)<br />
+  -&gt; shake at 37 degree over night<br />
+  <br />
+  '''07-07-2008'''<br />
+) Thin down overnight culture to OD~0.1<br />
+:10          -&gt;          0.66 = OD 660<br />
+  <br />
+µl                   =                  0.66<br />
+  x                          =                  0.1                        -&gt;   x = 758µl ~ 760µl in 50ml <br />
+   We mixed 760µl cell culture with 50ml DYT and shaked both at 37°C; Inoculation after 95min<br />
+<p>2) '''Inoculation of cell culture with M13mp18 phages'''<br />
+  ~ 50ml       cell culture<br />
+  + 5µl          M13mp18 phages (from -80°C)<br />
+  -&gt; 4h shaking at 37°C</p>
+<p>3) '''Phage precipitation'''<br />
+) centrifuging of the phage-solution at 5000 x g for 20 min.<br />
+) after decanting supernatant dessolve the pellet in 7ml (1/7 of the supernatant volume) PEG/NaCl<br />
+  -&gt; The precipitation occurs at 4°C over night</p>
 }}