User:Meng Xiao He/Notebook/fall08/2008/11/25

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Filter

 * cbb3, plate: >100 colonies, although sizes vary greatly, with many large
 * cbb3, liquid: MR-1 growth (from pellet color)
 * cco, liquid: looks like E. coli is growing? Pellet is white
 * Duo, plate: 2 colonies, medium sized
 * cbb3 and Duo, filter into liquid: both cultures thick, streaked for isolation on LB Gm plates
 * cco: no growth in filter+liquid yet, plated cultures had many (~50) white, colonies, that look like E. coli

Spin down

 * cbb3 streaked: scattered colonies at beginning of streak
 * cbb3, liquid: MR-1 growth (from pellet color)
 * cco, liquid: white pellet > E. coli?
 * Duo streaked: 2 colonies
 * plating cell mix: this seems like a back idea, as significant amounts of E. coli growth occurs in patches (not isolated colonies)

Colony PCR
Used Platinum Taq mix, 10min 94C initially cell boiling (maybe too much?)


 * No colony from conjugation gave bands, just smears as in first few lanes.
 * last 4 lanes: cbb3 WM plasmid miniprep PCR, Duo WM plasmid miniprep PCR, cco WM plasmid miniprep PCR
 * Duo doesn't seem to work at this annealing temp (56C)
 * colony PCR needs protocol change

Next

 * cco and new MR-1 grown up for new try w/ filter system
 * 2 colonies of Duo and cbb each grown up in LB, LB Gm, and on sucrose plate

Transformation update

 * Old transformations (last set that had no intervening nonselective growth period) developped yellow-orange contaminant colonies. Clearly not E. coli/MR-1 b/c growth was minimal overnight. Plates discarded.


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