Haynes Lab:Notebook/Engineering PC-TFs/2013/02/08: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
Line 6: | Line 6: | ||
| colspan="2"| | | colspan="2"| | ||
<!-- ##### 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. ##### --> | ||
== | ==2/8/2013== | ||
* * '''Perform BsmBI/ T4 ligase mediated assembly''' | * * '''Perform BsmBI/ T4 ligase mediated assembly''' | ||
* BsmBI cuts the DNA fragments and creates complementary overhangs. | * BsmBI cuts the DNA fragments and creates complementary overhangs. | ||
Line 59: | Line 59: | ||
| Total|| 10.0 || 10.0 | | Total|| 10.0 || 10.0 | ||
|} | |} | ||
<font size=3>'''Bacterial transformation'''</font> | |||
* Added total volume (10.0 μL) to 50 μL chemically competent cells (e.g., BL21) in a 2.0 mL tube. | |||
* Incubated on ice for 2 min., heat shock at 42°C for exactly 45 sec., and placed on ice. | |||
* Added 800 μL sterile SOC medium. | |||
* Grow with shaking at 37°C for 30 min.(''Taped tubes to the shaker rack.'') | |||
* Pelleted the cells at top speed in a microcentrifuge for 3 min. at room temperature. | |||
* Discarded the supernatant. Resuspended the cells in 100 μL LB + antibiotic. | |||
* Plate cells on pre-warmed LB agar + antibiotic. (''I plated 100 μL of the cells. Then put beads on plates and shook.) Protocol didn't specify.'') Grow overnight at 37°C. | |||
* Check results tomorrow. | |||
[[Image:plates_1_8_2013.jpg|400px||left|]] | |||
*''Still nothing. I'm going to start everything over and see if I can troubleshoot. Vi should be working on Type IIS Assembly this week. So I should have something going by the end of next week.'' | |||
Revision as of 13:13, 9 February 2013
Engineering PC-TFs | <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page <html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html> </html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html> | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
2/8/2013
|