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Revision as of 15:43, 25 August 2018
![](https://static.igem.org/mediawiki/2018/a/ab/T--Tec-Chihuahua--interlabLetters.png)
Interlab Study 2018
All of the 2018 iGEM teams were invited to participate in the Fifth International InterLaboratory Measurement Study in synthetic biology. Team Tec-Chihuahua is excited about working on an international and collaborative experiment!
The goal of the iGEM InterLab Study is to identify and correct the sources of systematic variability in synthetic biology measurements. In order to address this challenge, the iGEM committee provides researchers with detailed protocols and requires data analysis on the measurement of GFP. This year, the million dollar question is: Can we reduce lab-to-lab variability in fluorescence measurements by normalizing to absolute cell count or colony-forming units (CFUs) instead of OD?
For more information, check out iGEM's official InterLab Study webpage. Below are the protocols we used and the data that we collected for the InterLab Study.
Part I: Interlab Results
OD600 Reference point
LUDOX CL-X | H2O | |
---|---|---|
Replicate 1 | 0.054 | 0.04 |
Replicate 2 | 0.051 | 0.035 |
Replicate 3 | 0.051 | 0.035 |
Replicate 4 | 0.052 | 0.035 |
Arith. Mean | 0.052 | 0.036 |
Corrected Abs600 | 0.016 | |
Reference OD600 | 0.063 | |
OD600/Abs600 | 4 |
text
Particle Standard Curve
Number of Particles | 2.35E+08 | 1.18E+08 | 5.88E+07 | 2.94E+07 | 1.47E+07 | 7.35E+06 | 3.68E+06 | 1.84E+06 | 9.19E+05 | 4.60E+05 | 2.30E+05 | 0 |
---|---|---|---|---|---|---|---|---|---|---|---|---|
Replicate 1 | 0.601 | 0.236 | 0.171 | 0.105 | 0.071 | 0.053 | 0.045 | 0.039 | 0.038 | 0.037 | 0.036 | 0.035 |
Replicate 2 | 0.49 | 0.263 | 0.156 | 0.102 | 0.068 | 0.052 | 0.044 | 0.041 | 0.039 | 0.036 | 0.034 | 0.037 |
Replicate 3 | 0.519 | 0.269 | 0.191 | 0.098 | 0.07 | 0.051 | 0.043 | 0.04 | 0.037 | 0.039 | 0.037 | 0.034 |
Replicate 4 | 0.516 | 0.269 | 0.171 | 0.102 | 0.071 | 0.054 | 0.044 | 0.039 | 0.039 | 0.036 | 0.035 | 0.034 |
Arith. Mean | 0.532 | 0.259 | 0.172 | 0.102 | 0.07 | 0.053 | 0.044 | 0.04 | 0.038 | 0.037 | 0.036 | 0.035 |
Arith. Std Dev. | 0.048 | 0.016 | 0.014 | 0.003 | 0.001 | 0.001 | 0.001 | 0.001 | 0.001 | 0.001 | 0.001 | 0.001 |
Arith. Net Mean | 0.497 | 0.224 | 0.137 | 0.067 | 0.035 | 0.018 | 0.009 | 0.005 | 0.003 | 0.002 | 0.001 |
Particle Standard Curve
![](https://static.igem.org/mediawiki/2018/6/65/T--Tec-Chihuahua--Calibracion_2.1.png)
Particle Standard Curve (log scale)
![](https://static.igem.org/mediawiki/2018/c/c3/T--Tec-Chihuahua--Calibracion_2.2.png)
Number of Particles | 2.35E+08 | 1.18E+08 | 5.88E+07 | 2.94E+07 | 1.47E+07 | 7.35E+06 | 3.68E+06 | 1.84E+06 | 9.19E+05 | 4.60E+05 | 2.30E+05 |
---|---|---|---|---|---|---|---|---|---|---|---|
Mean Particles/Abs600 | 4.74E+08 | 5.25E+08 | 4.29E+08 | 4.41E+08 | 4.20E+08 | 4.20E+08 | 4.08E+08 | 3.87E+08 | 2.83E+08 | 2.30E+08 | 4.60E+08 |
Mean of med-high levels | 4.47E+08 |
Fluorescein uM | 10 | 5 | 2.5 | 1.25 | 0.625 | 0.313 | 0.156 | 0.078 | 0.039 | 0.0195 | 0.0098 | 0 |
---|---|---|---|---|---|---|---|---|---|---|---|---|
Replicate 1 | 1852 | 969.7 | 538.3 | 275.5 | 136.1 | 71.11 | 35.16 | 17.74 | 8.998 | 4.618 | 2.761 | 0.3263 |
Replicate 2 | 1929 | 956.8 | 523.7 | 284.7 | 132.6 | 66.21 | 36.66 | 17.95 | 8.892 | 4.856 | 2.553 | 0.34 |
Replicate 3 | 1922 | 977.1 | 516.5 | 281.1 | 150.3 | 75.65 | 40.22 | 19.54 | 10.57 | 5.299 | 2.931 | 0.3541 |
Replicate 4 | 1929 | 980.4 | 518.8 | 277.4 | 144.7 | 72.36 | 36.64 | 18.28 | 9.432 | 4.724 | 2.5 | 0.3667 |
Arith. Mean | 1.91E+03 | 9.71E+02 | 5.24E+02 | 2.80E+02 | 1.41E+02 | 7.13E+01 | 3.72E+01 | 1.84E+01 | 9.47E+00 | 4.87E+00 | 2.69E+00 | 3.47E-01 |
Arith. Std. Dev. | 3.75E+01 | 1.05E+01 | 9.79E+00 | 4.08E+00 | 8.06E+00 | 3.92E+00 | 2.15E+00 | 8.06E-01 | 7.68E-01 | 2.99E-01 | 1.98E-01 | 1.75E-02 |
Arith. Net Mean | 1.91E+03 | 9.71E+02 | 5.24E+02 | 2.79E+02 | 1.41E+02 | 7.10E+01 | 3.68E+01 | 1.80E+01 | 9.13E+00 | 4.53E+00 | 2.34E+00 |
Calibration 3.1
![](https://static.igem.org/mediawiki/2018/b/b5/T--Tec-Chihuahua--Calibracion_3.1.png)
Calibration 3.2
![](https://static.igem.org/mediawiki/2018/0/05/T--Tec-Chihuahua--Calibracion_3.2.png)
Fluorescein/a.u Measurements
Fluorescein uM | 10 | 5 | 2.5 | 1.25 | 0.63 | 0.31 | 0.16 | 0.08 | 0.04 | 0.02 | 0.01 |
---|---|---|---|---|---|---|---|---|---|---|---|
uM Fluorescein/a.u. | 5.24E-03 | 5.15E-03 | 4.77E-03 | 4.48E-03 | 4.45E-03 | 4.40E-03 | 4.24E-03 | 4.33E-03 | 4.28E-03 | 4.31E-03 | 4.17E-03 |
Mean uM Fluorescein/a.u. | 4.65E-03 | ||||||||||
MEFL/a.u. | 2.80E+10 |
Fluorescence Raw Readings
Hour 0 | Neg. Control | Pos. Control | Device 1 | Device 2 | Device 3 | Device 4 | Device 5 | Device 6 | LB + Chlor (blank) |
---|---|---|---|---|---|---|---|---|---|
Colony 1, Replicate 1 | 5.482 | 8.456 | 18.54 | 9.417 | 5.513 | 18.04 | 26.03 | 6.729 | 4.351 |
Colony 1, Replicate 2 | 4.915 | 8.344 | 18.3 | 9.125 | 5.328 | 17.97 | 25.61 | 6.249 | 4.244 |
Colony 1, Replicate 3 | 5.244 | 8.183 | 18.5 | 9.04v | 5.163 | 18.2v | 25.89 | 6.73 | 4.447 |
Colony 1, Replicate 4 | 5.426 | 8.369 | 17.98 | 8.748 | 5.434 | 17.18 | 25.73 | 6.637 | 4.321 |
Colony 2, Replicate 1 | 5.348 | 7.593 | 20.28 | 8.4 | 5.534 | 17.25 | 22.39 | 5.868 | 4.355 |
Colony 2, Replicate 2 | 5.211 | 7.304 | 18.93 | 8.121 | 5.333 | 16.32 | 21.94 | 5.796 | 4.402 |
Colony 2, Replicate 3 | 5.308 | 7.355 | 19.89 | 8.181 | 5.408 | 16.89 | 21.81 | 5.982 | 4.399 |
Colony 2, Replicate 4 | 5.376 | 7.487 | 19.38 | 8.325 | 5.272 | 16.78 | 21.2 | 5.992 | 4.458 |
Hour 6 | Neg. Control | Pos. Control | Device 1 | Device 2 | Device 3 | Device 4 | Device 5 | Device 6 | LB + Chlor (blank) |
---|---|---|---|---|---|---|---|---|---|
Colony 1, Replicate 1 | 5.216 | 11.56 | 45.15 | 8.42 | 5.249 | 37.7 | 44.03 | 7.702 | 2.608 |
Colony 1, Replicate 2 | 5.1 | 12.65 | 45.74 | 8.949 | 5.008 | 38.49 | 47.11 | 7.273 | 3.704 |
Colony 1, Replicate 3 | 5.051 | 12.33 | 45.71 | 14.25 | 5.063 | 37.05 | 46.05 | 7.627 | 3.382 |
Colony 1, Replicate 4 | 4.967 | 12.71 | 45.11 | 14.56 | 5.018 | 39.27 | 45.37 | 7.444 | 4.121 |
Colony 2, Replicate 1 | 4.917 | 12 | 46.24 | 11.25 | 5.11 | 40.01 | 41.62 | 6.062 | 3.163 |
Colony 2, Replicate 2 | 4.896 | 12.41 | 47.24 | 10.74 | 5.003 | 38.93 | 42.12 | 6.935 | 3.178 |
Colony 2, Replicate 3 | 4.973 | 12.03 | 47.16 | 10.94 | 5.224 | 40.58 | 40.07 | 6.718 | 3.464 |
Colony 2, Replicate 4 | 4.867 | 12.54 | 46.62 | 10.87 | 5.001 | 40.69 | 38.35 | 6.513 | 3.5 |
Abs600 Raw Readings
Hour 0 | Neg. Control | Pos. Control | Device 1 | Device 2 | Device 3 | Device 4 | Device 5 | Device 6 | LB + Chlor (blank) |
---|---|---|---|---|---|---|---|---|---|
Colony 1, Replicate 1 | 0.074 | 0.076 | 0.07 | 0.077 | 0.08 | 0.085 | 0.073 | 0.072 | 0.039 |
Colony 1, Replicate 2 | 0.073 | 0.079 | 0.07 | 0.072 | 0.073 | 0.075 | 0.073 | 0.074 | 0.041 |
Colony 1, Replicate 3 | 0.079 | 0.078 | 0.067 | 0.076 | 0.074 | 0.075 | 0.069 | 0.078 | 0.04 |
Colony 1, Replicate 4 | 0.077 | 0.082 | 0.073 | 0.077 | 0.079 | 0.072 | 0.071 | 0.077 | 0.04 |
Colony 2, Replicate 1 | 0.079 | 0.079 | 0.077 | 0.073 | 0.074 | 0.076 | 0.072 | 0.076 | 0.04 |
Colony 2, Replicate 2 | 0.079 | 0.075 | 0.072 | 0.07 | 0.071 | 0.069 | 0.07 | 0.071 | 0.04 |
Colony 2, Replicate 3 | 0.075 | 0.076 | 0.075 | 0.071 | 0.081 | 0.072 | 0.068 | 0.073 | 0.04 |
Colony 2, Replicate 4 | 0.074 | 0.076 | 0.073 | 0.067 | 0.073 | 0.076 | 0.093 | 0.073 | 0.039 |
Hour 6 | Neg. Control | Pos. Control | Device 1 | Device 2 | Device 3 | Device 4 | Device 5 | Device 6 | LB + Chlor (blank) |
---|---|---|---|---|---|---|---|---|---|
Colony 1, Replicate 1 | 0.176 | 0.155 | 0.187 | 0.113 | 0.177 | 0.16 | 0.122 | 0.152 | 0.037 |
Colony 1, Replicate 2 | 0.168 | 0.164 | 0.174 | 0.122 | 0.18 | 0.174 | 0.134 | 0.155 | 0.038 |
Colony 1, Replicate 3 | 0.169 | 0.154 | 0.166 | 0.174 | 0.182 | 0.161 | 0.129 | 0.154 | 0.037 |
Colony 1, Replicate 4 | 0.169 | 0.147 | 0.171 | 0.171 | 0.178 | 0.166 | 0.122 | 0.154 | 0.038 |
Colony 2, Replicate 1 | 0.172 | 0.175 | 0.173 | 0.163 | 0.18 | 0.185 | 0.129 | 0.136 | 0.038 |
Colony 2, Replicate 2 | 0.173 | 0.163 | 0.184 | 0.153 | 0.171 | 0.195 | 0.121 | 0.155 | 0.038 |
Colony 2, Replicate 3 | 0.178 | 0.165 | 0.178 | 0.166 | 0.185 | 0.208 | 0.132 | 0.148 | 0.038 |
Colony 2, Replicate 4 | 0.159 | 0.159 | 0.184 | 0.155 | 0.174 | 0.172 | 0.117 | 0.142 | 0.037 |
Part II: Colony Forming Units Results
Colony Counting for Dilution Factors Plates
Culture 1 | Culture 2 | ||||||
Replicate 1 | Replicate 2 | Replicate 3 | Replicate 1 | Replicate 2 | Replicate 3 | ||
Positive Control | 8.00E+04 | TNTC | TNTC | TNTC | TNTC | TNTC | TNTC |
8.00E+05 | 273 | TNTC | TNTC | 69 | 84 | 99 | |
8.00E+04 | 17 | TNTC | 41 | 9 | 35 | 20 | |
Negative Control | 8.00E+04 | TNTC | TNTC | TNTC | TNTC | TNTC | TNTC |
8.00E+05 | TNTC | TNTC | TNTC | TNTC | 283 | 300 | |
8.00E+04 | TNTC | 59 | 240 | 72 | 33 | 27 |
TNTC="Too Numerous To Count"
Dilution OD600 Measurements
Culture 1 | Culture 2 | |||||
Replicate 1 | Replicate 2 | Replicate 3 | Replicate 1 | Replicate 2 | Replicate 3 | |
Positive Control | 0.099 | 0.105 | 0.102 | 0.097 | 0.100 | 0.096 |
Negative Control | 0.109 | 0.103 | 0.105 | 0.101 | 0.101 | 0.099 |
CFU Calculations
Culture 1 | Culture 2 | |||||
Replicate 1 | Replicate 2 | Replicate 3 | Replicate 1 | Replicate 2 | Replicate 3 | |
Positive Control | 1.36E+08 | TNTC | 3.28E+08 | 5.52E+07 | 6.72E+07 | 7.92E+07 |
Negative Control | TNTC | 4.72E+08 | 1.92E+09 | 5.76E+08 | 2.64E+08 | 2.16E+08 |
TNTC="Too Numerous To Count"