Team:BU Wellesley Software/Notebook/AlbertoNotebook

From 2011.igem.org

2011 IGEM WetLab Notebook - Weekly Log

6/6/11-10/6/11

A. BFP2

Plasmid prep were created from the BFP2 culture plate and incubated for approximately 16 hours for plasmid amplification. The resulting plasmids were isolated by miniprep and quantified with nanodrop. The DNA concentration in the two samples are 16.5 ng/uL and 14.9 ng/uL, respectively. While such values are below the normally acceptable range of 20 and above, we still decided to run electrophoresis on them. No band was detected in the gel.

B. Pbad

Using the IGEM plate from spring 2010, Pbad promoter biobrick was obtained and transformed intoEe. coli bacteria. The bacteria were then plated onto agar-based LB+Ampicillin growth medium. Lots of cultures were found on the next day after the overnight transformation. However, they appeared to be either dark or light. Plasmid prep was done for each of the color type (pipet tip was used instead of metal loop) and each tube was incubated for 12 hours. On the next morning, the tip in each tube was found to contain a spiral, string-like thing and most of the plasmid preps were not opaque. After the miniprep, each of the plasmid was quantified with nanodrop. Their respective concentrations were 10.0 ng/uL and 8.3 ng/uL. They were found to be free from contaminants (based on the 260/280 and 260/230 values). But none of the samples appeared on the gel after electrophoresis.

In order to understand the problem that had been plaguing our work, we went to a biology lab to view the plasmid prep under compound microscope and conduct gram staining analysis. Apparently, all plasmid preps seemed to be contaminated with either another type of bacteria (due to differences in shape and size) or unidentified string-like compound. Many of the plasmid preps contain purple-colored bacteria, even though E. coli bacteria are supposed to be pink since they are gram-negative.