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Project

Lab Notebook / Lab Journal

Weeks 1 and 2 (June 1st - June 12th)

After many months of looking for a lab where we could work and after planning our project, we arrived to our lab with the Wetlab assesor and began sterilizing some material and locating many things we are going to use.

Also, our guests from the UNAH arrived and we were pleased to teach them some Molecular Biology and Protein’s Physicochemistry principles, as well as how to use scientific software and how to navigate in iGEM’s databases. They are staying in Mexico for 4 weeks.

Weeks 3 and 4 (June 15th - June 26th)

We went to Cuernavaca in order to speak with many researchers about our projects (ours and the one of our guests); we went to the Biotechnology Institute, the Center for Genomic Sciences and the Physics Institute.

We began some Wetlab practices with our UNAH guests and we inoculated the E. coli strains that we are going to use in our project. These strains are:

EnvZ -/- An E. coli strain with a deleterious mutation in the EnvZ receptor, which senses osmotic changes in the media and is part of the EnvZ/OmpR two component system.

SHuffle A strain with a cytoplasmic oxidative environment capable of making disulphide bonds in many peptides (which a WT strain is no able to form).

Rosettagami A strain used to synthesize many folded proteins which cannot be right folded by a WT strain.

DH5α Typical WT strain used for molecular biology steps like cloning vectors and amplifying plasmids. This strain methylates its DNA.

JM110 WT strain which doesn’t methylate its DNA, it is used mainly to obtain plasmids for digestion reactions.

All strains were cultured in LB medium

After we amplified our cultures, we were able to make competent cells with CaCl2 and we tried to transform EnvZ-/- and DH5α with our main reporter gene, RFP under OmpR promoter, but got only a single colony in DH5α strain, so we are going to make competent cells again.

Weeks 5 and 6 (June 29th - July 10th)

We followed DH5α, EnvZ-/-, SHuffle and Rosettagami’s growing rates by measuring absorbance at 600nm. They were cultured in complete LB medium.