Jessica Karen Wong/Notebook/2007-7-3

To Do

 * Colony PCR I2056 Tet
 * Replate I2056
 * Order new primer for I2055
 * Make overnights of other I2055 colonies for sequencing
 * Run gel of I2056 and T9002

I2055

 * Took out gradient PCR
 * Checked sequencing using AlignX and Finch TV on Jason's computer
 * I2055 had not ligated the R0040 to the I3401, aka the promoter was missing
 * We went back to the gel image to see if any other colony pcr's look a bit bigger and may include R0040
 * Made overnights of colonies 3,4, and 6 to miniprep for sequencing tomorrow
 * 5ml overnights were made with 5ul Amp and 5ul Kan b/c is on a 1AK3 plasmid
 * Also designed primers to PCR R0040 in if none of the colonies were successfully ligated
 * Only ordered a forward primer with Tail including Mfe1 and R0040
 * format: 8-mer.MfeI.promoter+mix.part of gfp (Tm)
 * CTTAGTAG.CAATTG.tccctatcagtgatagagattgacatccctatcagtgatagagatactgagcactactag.ATGCGTAAAGGAGAAGAACTTTTC (52.4)

I2056

 * Saw a colony on the Tet plate
 * Did a 10ul colony PCR on it, but result seemed slightly evaporated
 * Redoing the colony PCR overnight
 * Spun down and replated the rest of the transformants on both Tet and Chlor and are growing overnight

Gel

 * [[Image:7-3.jpg|thumb|left|Gel from 7/3/07]]
 * Ran a gel of the I2056 colony PCR and 10 ul of the T9002 100ul preparatory PCR
 * Loaded gel L to R: Ladder __ T9002 I2056 Ladder
 * Did a post stain with cybr gold
 * Scarred T9002 looks the right size
 * Redoing I2056

 2 log ladder