Difference between revisions of "Will Notebook2"
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| + | ==[[User:Wideloache|Wideloache]] 15:45, 15 January 2009 (EST)== | ||
| + | I ran my digestion of pSB1A2 (E/P) on a 0.8% gel and got the following result: | ||
| + | |||
| + | I expected to see a band at around 900, which would have been the RFP insert, but since the vector was fairly faint as well, it probably just wasn't bright enough to see. I excised the band at around 2000 bps which should be pSB1A2 vector cut with E/P. | ||
| + | |||
| + | I also ran my digestions of pLuxLacIX86 and pLuxLasLacII12X86 on a 2.0% gel and got the following result. | ||
| + | |||
| + | This was what I expected to see, and I excised the lowest band on each gel (expected size was 93 bps) | ||
| + | === === | ||
==[[User:Wideloache|Wideloache]] 20:36, 13 January 2009 (EST)== | ==[[User:Wideloache|Wideloache]] 20:36, 13 January 2009 (EST)== | ||
Today was my first day back in the lab of the semester. The first thing that I needed to do was to get the two pLux promoters that had been synthesized by GeneArt into pSB1A2. I had done an (E/P) digestion of pLux' and pLuxLas last semester that seemed to be successful. However, when I tried to transform into pSB1A2, I had a low colony yield. I decided to make a fresh stock of pSB1A2 digested with E/P. I used the part J61002-J23100. Cutting this with E/P woudl result in a pSB1A2 vector with a RFP expression insert. In this way I could screen out unsuccessful ligations. In addition to running this digestion O/N, I also redigested pLux' and pLuxLas with E/P from the minipreps of pLuxLacIX86 and pLuxLasLacII12X86. Tomorrow i will run these digestions next to my gel purifications from last semester to insure that those are the correct length insert. | Today was my first day back in the lab of the semester. The first thing that I needed to do was to get the two pLux promoters that had been synthesized by GeneArt into pSB1A2. I had done an (E/P) digestion of pLux' and pLuxLas last semester that seemed to be successful. However, when I tried to transform into pSB1A2, I had a low colony yield. I decided to make a fresh stock of pSB1A2 digested with E/P. I used the part J61002-J23100. Cutting this with E/P woudl result in a pSB1A2 vector with a RFP expression insert. In this way I could screen out unsuccessful ligations. In addition to running this digestion O/N, I also redigested pLux' and pLuxLas with E/P from the minipreps of pLuxLacIX86 and pLuxLasLacII12X86. Tomorrow i will run these digestions next to my gel purifications from last semester to insure that those are the correct length insert. | ||
Revision as of 20:45, 15 January 2009
Wideloache 15:45, 15 January 2009 (EST)
I ran my digestion of pSB1A2 (E/P) on a 0.8% gel and got the following result:
I expected to see a band at around 900, which would have been the RFP insert, but since the vector was fairly faint as well, it probably just wasn't bright enough to see. I excised the band at around 2000 bps which should be pSB1A2 vector cut with E/P.
I also ran my digestions of pLuxLacIX86 and pLuxLasLacII12X86 on a 2.0% gel and got the following result.
This was what I expected to see, and I excised the lowest band on each gel (expected size was 93 bps)
Wideloache 20:36, 13 January 2009 (EST)
Today was my first day back in the lab of the semester. The first thing that I needed to do was to get the two pLux promoters that had been synthesized by GeneArt into pSB1A2. I had done an (E/P) digestion of pLux' and pLuxLas last semester that seemed to be successful. However, when I tried to transform into pSB1A2, I had a low colony yield. I decided to make a fresh stock of pSB1A2 digested with E/P. I used the part J61002-J23100. Cutting this with E/P woudl result in a pSB1A2 vector with a RFP expression insert. In this way I could screen out unsuccessful ligations. In addition to running this digestion O/N, I also redigested pLux' and pLuxLas with E/P from the minipreps of pLuxLacIX86 and pLuxLasLacII12X86. Tomorrow i will run these digestions next to my gel purifications from last semester to insure that those are the correct length insert.
Make GenR plates (15 ug/mL) Miniprep pLas' and pLasLux Get GFP plasmid to put 4 promoters in front of.
