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Team:Calgary/23 June 2009
From 2009.igem.org
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| - | + | JUNE 23, 2009 | |
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| - | + | Multiple Miniprep isolations | |
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| - | + | * Attempted 15 miniprep reactions on both non-mutated and mutated <i>luxCDABE</i> plasmid. | |
| - | < | + | * The isolated plasmid was pooled together and attempted to concentrate the plasmid by spinning the tube in a vacifuge. After several hours (2.5 hours) the concentration was higher. |
| - | </ | + | * Tried ethanol precipitation earlier in the day but there was no precipitate found when done. |
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| - | + | Verification of NotI Digest | |
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| - | + | *Objective: To do one last verification that LuxOD47E is in the psB1AC3 vector before we send it down for DNA Sequecning. We will do a NotI digest with | |
| + | *Started with Plasmid Isolation this morning (Sigma). | ||
| + | *Used 1000 Nanodrop Spectrophotometer to determine concentrations. | ||
| + | *Set up a restriction digest with NotI enzyme and REact Buffer 3, left to digest in the 37 C waterbath overnight. | ||
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| - | + | Marketing for 23rd June 2009 | |
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| - | + | Today, I helped finish the July newsletter which will be ready be next week to sent out to our sponsors. Also I e-mailed some research and development companies and will do a follow-up soon. | |
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| - | + | Verify B0015-R0040 and luxOU-B0015 construction | |
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| + | [[Image:Calgary_luxOU_B0015-R0040+luxOU-B0015_NotI.jpg | 500px]] | ||
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| + | Figure. 1% agarose gel (100V) for NotI verification digest of construction yielding BBa_B0015-BBa_R0040 and luxOU(XbaI or EcoRI)-BBa_B0015. 200ng of plasmid was cut with NotI for 2 hours at 37oC. BBa_B0015-BBa_R0040 C1 is lane 2) and C4 is lane 4. luxOU(XbaI)-BBa_B0015 C3 is in lane 4 and C4 is in lane 5. luxOU(EcoRI)-BBa_B0015 C3 is in lane 6 and C4 in lane 7. Uncut luxOU in psB1AC3 (EcoRI C1 and XbaI C2) were used as positive controls in lanes 8 and 9, respectively. Expected band sizes for the BBa_B0015-BBa_R0040 construction are 3.2kb (psB1AK3) and 183bp. Expected band sizes for the luxOU-BBa_B0015 construction are 3kb (psB1AC3) and 2.1kb. 5μL of GeneRuler 1kb Plus DNA Ladder was loaded. | ||
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| + | BBa_B0015-BBa_R0040 C1 and luxOU(XbaI)-BBa_B0015 C4 were sent for sequencing with BBK-CP-F and BBK-CP-R as sequencing primers. Sequencing results were checked for vector contamination using “VecScreen”, protein sequences using “Blastx” and sequence conformation using “Blast2” (National Center for Biotechnology Information, MD). | ||
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| - | + | Isolating LuxPQ in psB1AK3 from overnights | |
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| - | + | Purpose: isolate LuxPQ in psB1AK3 from overnights of Colony 4 to continue construction of signaling circuit. Plasmid was isolated from colony 4 of luxPQ. This was done using the SIGMA’s Genelute plasmid Miniprep Kit. Plasmid purity and concentration were measured using the NanoDrop Spectrophotometer. | |
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Glycerol stocks of R0040, B0015 J13002, and Pqrr4 were made in order to preserve them long term. The addition of glycerl is important because otherwise the cellular membrane would shear. | Glycerol stocks of R0040, B0015 J13002, and Pqrr4 were made in order to preserve them long term. The addition of glycerl is important because otherwise the cellular membrane would shear. | ||
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| - | + | 2. Plasmid Isolation | |
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| - | + | To obtain more isolated R0040, B0015, J1302, and Pqrr4, miniprep (plasmid isolation) was done on them. | |
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| - | <a name=" | + | <a name="Mandy"></a> |
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| - | + | MANDY | |
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| - | + | Blog Making | |
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| - | + | Put together a workable blog and other online accounts for our team to update each other on what they've been up to, with an established schedule and everything :) Now I just have to figure out how to get it onto our wiki, or if I have to transfer each post manually from http://igemcalgary.blogspot.com | |
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| - | + | Marketing & Sponsorship | |
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| - | + | After constantly following up with companies over the past few weeks, I successfully recruited Alberta Research Council to sponsor the University of Calgary's iGEM team. They have generously donated %250 dollars this year. | |
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| + | We also edited the June newsletter and discussed it with the rest of the marketing team prior to sending it to all the companies we had contacted (including those who have declined our proposal). | ||
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| - | + | Sequencing of what I hope will be LuxOD47A BBK | |
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| - | + | I prepared a sample of the recently-formed-and-colony-PCR'd-and-NotI-restriction-digested LuxOD47A BBk for sequencing. This was done with BBk sequencing primers, in accordance with the sequencing protocol. | |
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Latest revision as of 06:26, 20 October 2009
UNIVERSITY OF CALGARY
