User:Anthony Salvagno/Notebook/Research/2009/03/16/DNA Digestion Day 3

From OpenWetWare

Jump to: navigation, search

Contents

Last Time

I ran a small gel of the pRS DNA and the gDNA and we found that XhoI hasn't been cutting so well. We decided to phenol extract and EtOH precipitate the remaining pure gDNA to clean it and then try again.

I ran the extraction and precipitation and today I did a nanodrop of the new clean stuff:
156.1ng/uL --> 23.4ug gDNA (in 150uL TE)
I used to have ~85ug gDNA. I've used a lot lately.

Digesting Again

We decided to run the digestion over night to try and get a good yield. I supposed expertise with these protocols would be really useful, so I can run a couple different methods side by side so that I can see which way gives the best cut.

Gel Prep
Tube Number Vol pRS413 Vol gDNA Vol and Type 10x Buffer Vol BSA 10x Vol XhoI Vol H2O Total Vol
2 ---- 32uL NE2/7.5uL 7.5uL 4uL 24uL 75uL
  • add 3uL XhoI after 2 hours.

Notes

I decided to keep the tube named number 2 even though I am only running one digest. The reason is because the pRSDNA is tube 1 and eventually we are going to do a ligation.

Annoyances

I had to get the image of my Friday gel off the computer. The computer attached to the UV lamp is literally from 1998. It is running Mac OS probably not X. It also has Netscape Navigator from waaaaaaaaaay back in the day and web pages take 5 hours to load (maybe at least 5 minutes). I try and go to sites that have as few images as possible because I believe the modem is 56k or something. I had to take a picture of the computer screen with my phone because the mac wouldn't even go to UNM's webmail which is as low tech as possible. That sucked.

I also had to fill out some ridiculously long IGERT survey which was ridiculously long. Not to be redundant, but did I mention how long it was? That wasted a good 40 minutes. Thank goodness digestions provide me with hours of downtime.

Another annoying thing is deal with the wireless internet on medical campus. At the Osley lab I barely get it, and it may even go out as I type this. I think there is a hard line, but I don't have my ethernet cable. Oh great it is out now... I have to wait for it to come back before I save this page. Hopefully it does before I get logged out of OWW... another annoyance.

Personal tools