Haynes Lab:Notebook/Engineering PC-TFs/2012/09/05: Difference between revisions
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<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### --> | <!-- ##### DO NOT edit above this line unless you know what you are doing. ##### --> | ||
==9/7/2012== | ==9/7/2012== | ||
{|border="1" cellpadding="5" cellspacing="0" align=" | {|border="1" cellpadding="5" cellspacing="0" align="right" | ||
|- | |||
! scope="col" style="background:#efefef;" | # | |||
! scope="col" style="background:#efefef;" | Biobrick | |||
! scope="col" style="background:#efefef;" | ng/µL | |||
! scope="col" style="background:#efefef;" | Size (bp) | |||
|- | |||
|1 | |||
|hPCD : mCh : SP1AB | |||
|130.56 | |||
|2520 | |||
|- | |||
|2 | |||
|hPCD : mCh : SP1A | |||
|266.898 | |||
|1443 | |||
|- | |||
|3 | |||
|hPCD : mCh : SP1B | |||
|222.175 | |||
|1224 | |||
|- | |||
|4 | |||
|hPCD : mCh : p65 | |||
|189.63 | |||
|1686 | |||
|- | |||
|5 | |||
|hPCD : mCh : VP64 | |||
|66.561 | |||
|1053 | |||
|- | |||
|6 | |||
|fshPCD : mCh : SP1AB | |||
|140.347 | |||
|2520 | |||
|- | |||
|7 | |||
|fshPCD : mCh : SP1A | |||
|148.285 | |||
|1443 | |||
|- | |||
|8 | |||
|fshPCD : mCh : SP1B | |||
|209.637 | |||
|1743 | |||
|- | |||
|9 | |||
|fshPCD : mCh : p65 | |||
|134.672 | |||
|1686 | |||
|- | |||
|10 | |||
|fshPCD : mCh : VP64 | |||
|263.567 | |||
|1053 | |||
|- | |||
|11 | |||
|flyPCD : mCh : SP1A | |||
|43.068 | |||
|1443 | |||
|- | |||
|12 | |||
|flyPCD : mCh : SP1B | |||
|35.838 | |||
|1743 | |||
|- | |||
|13 | |||
|flyPCD : mCh : p65 | |||
|83.886 | |||
|1686 | |||
|- | |||
|14 | |||
|flyPCD : mCh : VP64 | |||
|200.896 | |||
|1053 | |||
|- | |||
|} | |||
<br> | |||
{|border="1" cellpadding="5" cellspacing="0" align="right" | |||
|- | |- | ||
! scope="col" style="background:#efefef;" | Ligation | ! scope="col" style="background:#efefef;" | Ligation | ||
Line 60: | Line 139: | ||
|.3 μL | |.3 μL | ||
|- | |- | ||
|T4 Ligase | |T4 Ligase | ||
|1 μL | |||
|1 μL | |1 μL | ||
|1 μL | |1 μL | ||
Line 94: | Line 173: | ||
|5 μL | |5 μL | ||
|- | |- | ||
|dH2O | |dH2O | ||
|3.4 μL | |3.4 μL | ||
Line 112: | Line 190: | ||
|3.7 μL | |3.7 μL | ||
|- | |- | ||
| | |Total | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
| | |10 μL | ||
|10 μL | |||
|10 | |||
|- | |- | ||
|} | |} | ||
*In .5 ml tubes, pipette the ligation buffer first. | |||
*Then pipette water, DNA insert, DNA vector, and T4 ligase in this order. | |||
*Allowed them to incubate for 10 minutes. | |||
*Retrieved ice bucket and allowed E.coli cells to thaw for five minutes. | |||
*Followed the transformation process: | |||
*Warmed 100 agar plates at 37°C. | |||
*Thaw two fresh tubes of DH5α Turbo cells on ice. | |||
*Add 30 μL of DH5α Turbo cells to DNA | |||
*Incubated cells + DNA on ice for 5-10 minutes. | |||
*Transfered cells + DNA onto agar plates | |||
*Added 10-15 sterile glass beads to plates and shook plates. | |||
*Discarded beads and incubated plates at 37°C°C overnight. | |||
Revision as of 18:35, 7 September 2012
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9/7/2012
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