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Team:Buenos Aires/Results/Strains
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Through our experiments we worked with the following strains: | Through our experiments we worked with the following strains: | ||
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Nearly 15 other similar strains were evaluated and discarded due to several reasons (low screening potentiality; requirement of hormones for fluorescence induction; high reverting rate of auxotrophies, among others) | Nearly 15 other similar strains were evaluated and discarded due to several reasons (low screening potentiality; requirement of hormones for fluorescence induction; high reverting rate of auxotrophies, among others) | ||
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== b) Fluorescence screening == | == b) Fluorescence screening == | ||
Revision as of 00:57, 16 September 2012
Contents |
Strain characterization
a) Description of strains
Through our experiments we worked with the following strains:
| Strain ID | Relevant Auxotrophies | Fluorescence | Used as |
| TCY 3043 | (-H-T) | No fluorescence | Negative control |
| TCY 3190 | (+H-T) | YFP + (Induced CFP) | For coculture |
| TCY 3265 | (-H+T) | CFP | For coculture |
| TCY 3154 | CFP +(induced YFP) | Positive Control |
Table: Hystidine (H) and Tryptophane (T) auxotrophies per strain, type of fluorescence and description of most common utilization during the experiments.
Nearly 15 other similar strains were evaluated and discarded due to several reasons (low screening potentiality; requirement of hormones for fluorescence induction; high reverting rate of auxotrophies, among others)
Foto 1
b) Fluorescence screening
We measured Strains 3281 (YFP) and 3265 (CFP) and got a spectrum of each one prooving that these strains can be distinguished by their fluorescence in culture.
Graficos 1
c) Auxotrophy confirmation
Several times during the experiments we control and checked if the auxotrophies in the selected strain were functional by plating all of them in medium deficient in aminoacids (-H; -T; -H-T and control +H+T). We observed differential growth according to expected due to the description of each strain in point a)
Foto 2,3,4,5
Figures a-d: Auxotrophies check. Plate a : medium complete. Plate b: medium without T. Plate c: medium without H and T. Plate d: medium without H.
We observed all the strains grew in Plate a and only 3154 (+H+T) grew in Plate c. In plate B, only those strains able to synthesize T grew (3265 and 3154) and in Plate d, only those able to produce H grew (3190 and 3154), as expected. This means our strains work according to their description. We did this several times during the months to check for reversions or contaminations.
d) Coculture in liquid medium: growth
| Coculture | Proportion (H+T-):(H-T+) | Initial OD(t=0) | OD1 (t=1) | OD2 (t=1) | dilution used for measure t=1 | Mean OD | Growth Factor |
| 01:01 | 0,25 | 0,32 | 0,314 | 10 | 3,17 | 12,68 | |
| 09:01 | 0,25 | 0,148 | 0,144 | 10 | 1,46 | 5,84 | |
| 01:09 | 0,25 | 0,138 | 0,189 | 10 | 1,635 | 6,54 | |
| 01:01 | 0,1 | 0,109 | 0,169 | 10 | 1,39 | 13,9 | |
| 09:01 | 0,1 | 0,04 | 0,045 | 10 | 0,425 | 4,25 | |
| 01:09 | 0,1 | 0,067 | 0,053 | 10 | 0,6 | 6 | |
| 01:01 | 0,01 | 0,067 | 0,061 | 1 | 0,064 | 6,4 | |
| 09:01 | 0,01 | 0,056 | 0,05 | 1 | 0,053 | 5,3 | |
| 01:09 | 0,01 | 0,074 | 0,073 | 1 | 0,0735 | 7,35 |
Table: Coculture in at different initial OD and proportions (Days 0 and 1)
Graficos 2,3,4
As shown in graphs there is a basal growth that does not depend on the initial OD or strain proportion, of a growth factor of 6 approximately. However we observed a much higher growth at the proportion 1:1 when the initial OD 0.25 and 0.1. Therefore we can assume that at these proportions there is a natural cooperation between the strains and that should be the level of growth that we would like to assess through our bioengineering. Besides we would like to be able in the future to tune the strains in order to be able to obtain in the proportions 9:1 and 1:9 similar results to those obtained in the 1:1, at our own will.
