iGEM 2016 — Team Ryerson_Toronto

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# Team:Ryerson Toronto/Notebook

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iGEM Ryerson University 2016

# Notebook

### Week 1

#### P. areuginosa, strain mPA01, grown on LB agar plate overnight at 37°C

![](https://static.igem.org/mediawiki/2016/f/fe/T--Ryerson_Toronto--RUpseudomon.jpg)

Gentamycin, at a final concentration of 30μg/mL-100μg/mL, was used to counter select

#### Single colony grown overnight in LB broth

#### Genomic DNA extracted

LB broth was spun down, resuspended in PBS, and boiled for 20 min. The supernatant was then transferred to a new tube

### Week 2

#### PCR

samples were prepared in GC and HF Phusion buffer master mixes, with the following sample dilutions: no dilution, 1:50, 1:100, 1:250, 1:1000

#### Gel Electrophoresis

agarose gel, at a final concentration of 0.8% made in TBE, was run at 100V for 60min

### Week 3

#### Transformation

For restriction digestion for transformation, the following enzymes were used: kpni and xhol

Samples from Transformation were applied to a gel

1% agarose gel was made with TAE, and run at 40V for 60min

#### T4 ligation

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