Difference between revisions of "Team:Reading/Notebook"
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<title2> <b> 23/07/15 </b> </title2> | <title2> <b> 23/07/15 </b> </title2> |
Revision as of 11:38, 18 September 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.
Stock growth onto LB kan 50 plates unsuccessful.
Potassium ferricyanide reagent arrives, protocols start being drawn up
Potassium ferricyanide reagent arrives, protocols start being drawn up
Nanowires concluded to be a non-fruitful venture
Our lab becomes engulfed in the tides of cuvettes arriving
Our lab becomes engulfed in the tides of cuvettes arriving
Competition assay starting to reach log phase
LB BG-11 Kan plates and LB BG-11 plates made up to plate the competition assay on
Competition assay streaked onto plates