Difference between revisions of "Team:Reading/Notebook"
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<title1> Lab notes</title1> | <title1> Lab notes</title1> | ||
<p> This page is an account of the daily events in our lab and project. Dates are in the UK format DD/MM/YY. 6803 refers to the cyanobacterium used - <i>Synechocystis sp. PCC 6803</i> </p> | <p> This page is an account of the daily events in our lab and project. Dates are in the UK format DD/MM/YY. 6803 refers to the cyanobacterium used - <i>Synechocystis sp. PCC 6803</i> </p> | ||
| − | < | + | <title2> <b> 29/06/15 </b> </title2> |
<li> Cultured a sterile 500ml conical flask using 100 ml BG-11 and 5 loopfuls of <i>6803</i> Wild Type (WT). (Flask 1) </li> | <li> Cultured a sterile 500ml conical flask using 100 ml BG-11 and 5 loopfuls of <i>6803</i> Wild Type (WT). (Flask 1) </li> | ||
<li> Cultured 5 streak plates of <i>6803</i> WT on BG-11 Agar </li> | <li> Cultured 5 streak plates of <i>6803</i> WT on BG-11 Agar </li> | ||
<p> All placed on the windowsill in our lab under natural light and at room temperature (~25°C) </p> | <p> All placed on the windowsill in our lab under natural light and at room temperature (~25°C) </p> | ||
<!-- Pete please Insert image of flask and plates on windowsill --> | <!-- Pete please Insert image of flask and plates on windowsill --> | ||
| − | < | + | <title2> <b> 02/07/15 </b> </title2> |
<li> Three new sterile 500ml conical flasks set up on the shaker at 58rpm and 30°C. Each contains 5 loopfuls of <i>6803</i> WT and 100ml of BG-11. They are exposed to the overhead lights in the lab 24/7. (Flasks 2, 3, 4) </li> | <li> Three new sterile 500ml conical flasks set up on the shaker at 58rpm and 30°C. Each contains 5 loopfuls of <i>6803</i> WT and 100ml of BG-11. They are exposed to the overhead lights in the lab 24/7. (Flasks 2, 3, 4) </li> | ||
| − | < | + | <title2> <b> 07/07/15 </b> </title2> |
<li>Flask 5 cultured containing 10ul <i>6803</i> WT in 100ml BG-11. This flask will be used for preliminary growth curve measurements. The flask was placed on the shaker with flasks 2,3 and 4 all at 58rpm and 30°C under lab overhead lights. </li> | <li>Flask 5 cultured containing 10ul <i>6803</i> WT in 100ml BG-11. This flask will be used for preliminary growth curve measurements. The flask was placed on the shaker with flasks 2,3 and 4 all at 58rpm and 30°C under lab overhead lights. </li> | ||
| − | < | + | <title2> <b> 10/07/15 </b> </title2> |
<p> 2 Litres of water collected from Whiteknights Lake on campus for SODIS, method outlines in protocols. </p> | <p> 2 Litres of water collected from Whiteknights Lake on campus for SODIS, method outlines in protocols. </p> | ||
| − | < | + | <title2> <b> 13/07/15 </b> </title2> |
<p> Estimating the Cells per ml for the flask from 29/06 1 a.u at 750nm = 1.6x10<sup>8</sup> <li> OD<sub>750</sub> = 0.028 </li> Therefore <li> 0.028x1.6x10<sup>8</sup> = 4.48x10<sup>6</sup> cells per ml </li> </p> | <p> Estimating the Cells per ml for the flask from 29/06 1 a.u at 750nm = 1.6x10<sup>8</sup> <li> OD<sub>750</sub> = 0.028 </li> Therefore <li> 0.028x1.6x10<sup>8</sup> = 4.48x10<sup>6</sup> cells per ml </li> </p> | ||
<p> OD<sub>750</sub> measurement and cells per ml estimated for flasks cultured on 02/07 </p> | <p> OD<sub>750</sub> measurement and cells per ml estimated for flasks cultured on 02/07 </p> | ||
| − | < | + | <title2> <b> 14/07/15 </b> </title2> |
<p> The lamp safety testing completed so flasks placed under the bright white light bulb </p> | <p> The lamp safety testing completed so flasks placed under the bright white light bulb </p> | ||
| − | < | + | <title2> <b> 21/07/15 </b> </title2> |
<p> Flasks are getting very green, definitely a good sign! </p> | <p> Flasks are getting very green, definitely a good sign! </p> | ||
| − | < | + | <title2> <b> 23/07/15 </b> </title2> |
<p> Flask 2 discarded as it is not showing growth, and has signs of contamination. We received most of the lab equipment we have ordered. </p> | <p> Flask 2 discarded as it is not showing growth, and has signs of contamination. We received most of the lab equipment we have ordered. </p> | ||
| − | < | + | <title2> <b> 27/07/15 </b> </title2> |
BG-11 order arrived finally! | BG-11 order arrived finally! | ||
<ol> | <ol> | ||
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</ol> | </ol> | ||
<p> All flasks cultured today were placed in the incubator, being shaken at 58rpm, 30°C, and under bright white light </p> | <p> All flasks cultured today were placed in the incubator, being shaken at 58rpm, 30°C, and under bright white light </p> | ||
| − | < | + | <title2> <b> 05/08/15 </b> </title2> |
<p> 200ml of BG-11 was added to flasks 3 and 4 which already contained ~100ml of ~2 a.u density 6803 WT. </p> | <p> 200ml of BG-11 was added to flasks 3 and 4 which already contained ~100ml of ~2 a.u density 6803 WT. </p> | ||
<p> Competition Assay </p> | <p> Competition Assay </p> | ||
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<p> added 10ul of WT 6803 and 10ul of MT3 6803 </p> | <p> added 10ul of WT 6803 and 10ul of MT3 6803 </p> | ||
<p> Flask was placed in the incubator, being shaken at 58rpm, 30°C, and under bright white light </p> | <p> Flask was placed in the incubator, being shaken at 58rpm, 30°C, and under bright white light </p> | ||
| − | < | + | <title2> <b> 06/08/15 </b> </title2> |
<p> DNA concentration from miniprep </p> | <p> DNA concentration from miniprep </p> | ||
<ul> | <ul> | ||
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</ul> | </ul> | ||
<p> *no DNA due to miniprep from Glycerol Stock </p> | <p> *no DNA due to miniprep from Glycerol Stock </p> | ||
| − | < | + | <title2> <b> 07/08/15 </b> </title2> |
<p> Made LB Agar tranplates for culturing </p> | <p> Made LB Agar tranplates for culturing </p> | ||
<ul> | <ul> | ||
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<li>PilT</li> | <li>PilT</li> | ||
</ul> | </ul> | ||
| − | < | + | <title2> <b> 10/08/15 </b> </title2> |
<p> No growth on plates from 07/08 </p> | <p> No growth on plates from 07/08 </p> | ||
<p> Cultured plates of LB Agar from all stocks (glycerol stocks) </p> | <p> Cultured plates of LB Agar from all stocks (glycerol stocks) </p> | ||
| − | < | + | <title2> <b> 11/08/15 </b> </title2> |
<p> Growth on many the stocks from 10/08, replated onto LB kan 50 plates </p> | <p> Growth on many the stocks from 10/08, replated onto LB kan 50 plates </p> | ||
| − | < | + | <title2> <b> 12/08/15 </b> </title2> |
<p> Competition Assay had begun to turn a light green colour. </p> | <p> Competition Assay had begun to turn a light green colour. </p> | ||
Revision as of 10:54, 27 August 2015
This page is an account of the daily events in our lab and project. Dates are in the UK format DD/MM/YY. 6803 refers to the cyanobacterium used - Synechocystis sp. PCC 6803
All placed on the windowsill in our lab under natural light and at room temperature (~25°C)
2 Litres of water collected from Whiteknights Lake on campus for SODIS, method outlines in protocols.
Estimating the Cells per ml for the flask from 29/06 1 a.u at 750nm = 1.6x108
OD750 measurement and cells per ml estimated for flasks cultured on 02/07
The lamp safety testing completed so flasks placed under the bright white light bulb
Flasks are getting very green, definitely a good sign!
Flask 2 discarded as it is not showing growth, and has signs of contamination. We received most of the lab equipment we have ordered.
Miniprep of plasmid DNA: We carried out extraction of biobrick construct plasmids from E.coli NEB-5α glycerol stocks.
We extracted pSB1K3 plasmid backbones combined with biobricks containing the following genes:
- PilA1
- PsaD
- PetF
- PilT
This gave us 4 samples of plasmid DNA, which were placed in the freezer at -20°C.
-
BG-11 cultures: We added ~200ml of BG-11 to two sterile 500ml conical flasks. We inoculated them as follows;
- Flask 5 - 1ml of Wild type 6803 from flask 3 as this flask had the best growth.
- Flask 6 - 10µl of stock Howe Lab triple mutant 6803.
- SODIS lakewater cultures: We added ~200ml of SODIS lakewater to three sterile 500ml conical flasks. We inoculated them as follows;
- SODIS 1 and 2 - 1ml of Wild type 6803 from flask 3 as this flask had the best growth.
- SODIS 3 - 1ml of stock Howe Lab triple mutant 6803.
All flasks cultured today were placed in the incubator, being shaken at 58rpm, 30°C, and under bright white light
200ml of BG-11 was added to flasks 3 and 4 which already contained ~100ml of ~2 a.u density 6803 WT.
Competition Assay
200ml of BG-11 added to a new sterile conical flask
added 10ul of WT 6803 and 10ul of MT3 6803
Flask was placed in the incubator, being shaken at 58rpm, 30°C, and under bright white light
DNA concentration from miniprep
- PilA1 = 11.16ns/ul
- PsaD = 13.01ns/ul
- PetF = 0.38ns/ul
- PilT = 8.58ns/ul
*no DNA due to miniprep from Glycerol Stock
Made LB Agar tranplates for culturing
- PilA1
- PsaD
- PetF
- PilT
No growth on plates from 07/08
Cultured plates of LB Agar from all stocks (glycerol stocks)
Growth on many the stocks from 10/08, replated onto LB kan 50 plates
Competition Assay had begun to turn a light green colour.