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Team:Freiburg Bioware/NoteBook/Labjournal/October2
From 2010.igem.org
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(→cloning of lITR_CMV_betaglobin and lITR_phTERT_betaglobin into pSB1C3_CD) |
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====<p style="font-size:15px; background-color:#66bbff;"><b>cloning of lITR_CMV_betaglobin and lITR_phTERT_betaglobin into pSB1C3_CD</b></p>==== | ====<p style="font-size:15px; background-color:#66bbff;"><b>cloning of lITR_CMV_betaglobin and lITR_phTERT_betaglobin into pSB1C3_CD</b></p>==== | ||
| - | <b>Investigator: Stefan</b> | + | <b>Investigator: Stefan</b><br> |
| + | |||
| + | <p style="font-size:13px; color:red;">Comment: In order to test our GOI with another promotor, lITR_phTERT_beta-globin_mGMK_TK30 needs to be cloned upstream of hgH_rITR. </p><br /> | ||
| + | |||
| + | Vector name:<br /> | ||
| + | pSB1C3_CD clone 1 (P???)<br /> | ||
| + | pSB1C3_CD clone 2 (P???)<br /> | ||
| + | |||
| + | Insert name:<br /> | ||
| + | pSB1C3_lITR_CMV_beta-globin (P729)<br /> | ||
| + | pSB1C3_lITR_phTERT_beta-globin (P730)<br /> | ||
| + | |||
| + | <b>Digestion:</b><br /><br /> | ||
| + | |||
| + | <br /> | ||
| + | {| border="1" | ||
| + | | '''components''' || align="right" |'''volume CD clone 1 + 2 /µl'''|| align="right" |'''volume P729 /µl''''''volume P730 /µl''' | ||
| + | |- | ||
| + | | DNA || align="right" |6 || align="right" |14|| align="right" |6 | ||
| + | |- | ||
| + | | BSA (10x) || align="right" |2 || align="right" |2|| align="right" |2 | ||
| + | |- | ||
| + | | Buffer 4 (10x)|| align="right" |2 || align="right" |2|| align="right" |2 | ||
| + | |- | ||
| + | |Enzyme EcoI|| align="right" |1|| align="right" |1 || align="right" |1 | ||
| + | |- | ||
| + | |Enzyme XbaI|| align="right" |1|| align="right" |- || align="right" |- | ||
| + | |- | ||
| + | |Enzyme SpeI|| align="right" |-|| align="right" |1|| align="right" |1 | ||
| + | |- | ||
| + | |H2O|| align="right" |8|| align="right" |- || align="right" |8 | ||
| + | |- | ||
| + | |'''Total volume (e.g. 15,20,25,30 µl)'''|| align="right" | 20|| align="right" | 20|| align="right" | 20 | ||
| + | |} | ||
| + | |||
| + | <br /> | ||
| + | <b>Gel:</b><br /> | ||
| + | 0,5 g Agarose, 50 ml TAE (1%), 3 µl GELRED , at 115 Volt<br /> | ||
| + | |||
| + | |||
| + | <br/> | ||
| + | [[Image:new.jpg|550px|]]<br/> | ||
| + | |||
| + | <br/> | ||
| + | |||
| + | |||
| + | <b>Gel extraction</b>: <br> | ||
| + | Was performed according to protocol. | ||
| + | |||
| + | <br> | ||
| + | <b>T4 Ligation</b>: <br> | ||
| + | {| border="1" | ||
| + | |ligation name || align="right" |728 + 670 | ||
| + | |- | ||
| + | |volume of vector || align="right" |2,35 | ||
| + | |- | ||
| + | |volume of insert|| align="right" |5,65 | ||
| + | |- | ||
| + | |T4 ligase buffer (10x)|| align="right" |1 | ||
| + | |- | ||
| + | |T4 ligase || align="right" |1 | ||
| + | |- | ||
| + | |} | ||
| + | |||
| + | <br> | ||
| + | <b>Transformation</b>: <br> | ||
| + | Was performed according to standard protocol using BL21 cells. | ||
| + | <br/> | ||
===<p style="font-size:17px; background-color:#00dd77;">147. labday 12.10.2010: Example Example Example Example Example</p>=== | ===<p style="font-size:17px; background-color:#00dd77;">147. labday 12.10.2010: Example Example Example Example Example</p>=== | ||
Revision as of 11:48, 11 October 2010
- March (labday 1)
- April (labday 2 - 5)
- May (labday 6 - 17)
- June (labday 18 - 45)
- July (labday 46 - 75)
- August part 1 (labday 76 - 92)
- August part 2 (labday 93 - 106)
- September part 1 (labday 107 - 123)
- September part 2 (labday 124 - 135)
- October part 1 (labday 136 - 145 )
- October part 2 (labday 146 - 155 )
- October part 3 (labday 156 - 166 )
- November (labday 167 - 170 )
- Cellculture
Contents |
146. labday 11.10.2010
cloning of lITR_CMV_betaglobin and lITR_phTERT_betaglobin into pSB1C3_CD
Investigator: Stefan
Comment: In order to test our GOI with another promotor, lITR_phTERT_beta-globin_mGMK_TK30 needs to be cloned upstream of hgH_rITR.
Vector name:
pSB1C3_CD clone 1 (P???)
pSB1C3_CD clone 2 (P???)
Insert name:
pSB1C3_lITR_CMV_beta-globin (P729)
pSB1C3_lITR_phTERT_beta-globin (P730)
Digestion:
| components' | volume CD clone 1 + 2 /µl | volume P729 /µl'volume P730 /µl | |
| DNA | 6 | 14 | 6 |
| BSA (10x) | 2 | 2 | 2 |
| Buffer 4 (10x) | 2 | 2 | 2 |
| Enzyme EcoI | 1 | 1 | 1 |
| Enzyme XbaI | 1 | - | - |
| Enzyme SpeI | - | 1 | 1 |
| H2O | 8 | - | 8 |
| Total volume (e.g. 15,20,25,30 µl) | 20 | 20 | 20 |
Gel:
0,5 g Agarose, 50 ml TAE (1%), 3 µl GELRED , at 115 Volt
Gel extraction:
Was performed according to protocol.
T4 Ligation:
| ligation name | 728 + 670 |
| volume of vector | 2,35 |
| volume of insert | 5,65 |
| T4 ligase buffer (10x) | 1 |
| T4 ligase | 1 |
Transformation:
Was performed according to standard protocol using BL21 cells.
147. labday 12.10.2010: Example Example Example Example Example
148. labday 13.10.2010
149. labday 14.10.2010
150. labday 15.10.2010
151. labday 16.10.2010
152. labday 17.10.2010
153. labday 18.10.2010
154. labday 19.10.2010
155. labday 20.10.2010
