<p><b>Figure 1. FITC standard curve.</b> There is an approximate linear trend for increased fluorescence as [FITC] increases.</p>
+
</figure>
+
<figure>
+
<div id="container-2"></div>
+
<p><b>Figure 2. FITC standard curve (log scale).</b> After changing the curve into log scale, the linear trend becomes more remarkable and reliable.</p>
+
</figure>
+
<figure>
+
<table>
+
<thead>
+
<tr>
+
<th>Unit Scaling Factors:</th>
+
<th></th>
+
</tr>
+
</thead>
+
<tbody>
+
<tr>
+
<th>OD600/Abs600</th>
+
<td class="td-highlight">11.58</td>
+
</tr>
+
<tr>
+
<th>uM Fluorescein/a.u.</th>
+
<td class="td-highlight">0.003458</td>
+
</tr>
+
</tbody>
+
</table>
+
<p><b>Table 4. Normalization of initial concentration of colony.</b> In order to unify the initial concentration before , we use Varioskan Flash to measure the OD600 after incubating overnight.</p>
<p class="mdc-typography--body2">Because of the large account of data, we only use an average data in this form. (Average of 4 replicates of one colony)</p>
+
<figure>
+
<table>
+
<thead>
+
<tr>
+
<th>Abs600</th>
+
<th>Replicate</th>
+
<th>0h</th>
+
<th>2h</th>
+
<th>4h</th>
+
<th>6h</th>
+
</tr>
+
</thead>
+
<tbody>
+
<tr>
+
<th rowspan="2">Negative control</th>
+
<th>Colony 1</th>
+
<td>0.0666</td>
+
<td>0.1893</td>
+
<td>0.2708</td>
+
<td>0.4942</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0615</td>
+
<td>0.1929</td>
+
<td>0.3000</td>
+
<td>0.5681</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Positive Control</th>
+
<th>Colony 1</th>
+
<td>0.0601</td>
+
<td>0.1638</td>
+
<td>0.2949</td>
+
<td>0.5425</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0613</td>
+
<td>0.1839</td>
+
<td>0.2982</td>
+
<td>0.5815</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device1</th>
+
<th>Colony 1</th>
+
<td>0.0647</td>
+
<td>0.0649</td>
+
<td>0.0757</td>
+
<td>0.0816</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0560</td>
+
<td>0.0612</td>
+
<td>0.0627</td>
+
<td>0.0732</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device2</th>
+
<th>Colony 1</th>
+
<td>0.0616</td>
+
<td>0.1685</td>
+
<td>0.2480</td>
+
<td>0.4316</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0607</td>
+
<td>0.1928</td>
+
<td>0.2977</td>
+
<td>0.4638</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device3</th>
+
<th>Colony 1</th>
+
<td>0.0463</td>
+
<td>0.1566</td>
+
<td>0.3120</td>
+
<td>0.4887</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0521</td>
+
<td>0.1291</td>
+
<td>0.3464</td>
+
<td>0.6376</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device4</th>
+
<th>Colony 1</th>
+
<td>0.0571</td>
+
<td>0.1329</td>
+
<td>0.2899</td>
+
<td>0.4263</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0551</td>
+
<td>0.1259</td>
+
<td>0.2656</td>
+
<td>0.5216</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device5</th>
+
<th>Colony 1</th>
+
<td>0.0528</td>
+
<td>0.1776</td>
+
<td>0.3140</td>
+
<td>0.5123</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0655</td>
+
<td>0.2286</td>
+
<td>0.3704</td>
+
<td>0.5788</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device6</th>
+
<th>Colony 1</th>
+
<td>0.0575</td>
+
<td>0.1316</td>
+
<td>0.2915</td>
+
<td>0.5249</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0536</td>
+
<td>0.1530</td>
+
<td>0.3032</td>
+
<td>0.5852</td>
+
</tr>
+
</tbody>
+
</table>
+
<p><b>Tablet5. Raw Abs600 measurement.</b></p>
+
</figure>
+
+
<figure>
+
<table>
+
<thead>
+
<tr>
+
<th>Abs600</th>
+
<th>Replicate</th>
+
<th>0h</th>
+
<th>2h</th>
+
<th>4h</th>
+
<th>6h</th>
+
</tr>
+
</thead>
+
<tbody>
+
<tr>
+
<th rowspan="2">Negative control</th>
+
<th>Colony 1</th>
+
<td>0.0256</td>
+
<td>0.1483</td>
+
<td>0.2298</td>
+
<td>0.4942</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0205</td>
+
<td>0.1518</td>
+
<td>0.2590</td>
+
<td>0.5271</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Positive Control</th>
+
<th>Colony 1</th>
+
<td>0.0191</td>
+
<td>0.1227</td>
+
<td>0.2539</td>
+
<td>0.5014</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0203</td>
+
<td>0.1429</td>
+
<td>0.2571</td>
+
<td>0.5404</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device1</th>
+
<th>Colony 1</th>
+
<td>0.0237</td>
+
<td>0.0239</td>
+
<td>0.0347</td>
+
<td>0.0405</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0149</td>
+
<td>0.0202</td>
+
<td>0.0217</td>
+
<td>0.0321</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device2</th>
+
<th>Colony 1</th>
+
<td>0.0206</td>
+
<td>0.1274</td>
+
<td>0.2069</td>
+
<td>0.3905</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0197</td>
+
<td>0.1518</td>
+
<td>0.2567</td>
+
<td>0.4228</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device3</th>
+
<th>Colony 1</th>
+
<td>0.0053</td>
+
<td>0.1156</td>
+
<td>0.2710</td>
+
<td>0.4477</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0110</td>
+
<td>0.0881</td>
+
<td>0.3053</td>
+
<td>0.5966</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device4</th>
+
<th>Colony 1</th>
+
<td>0.0161</td>
+
<td>0.0919</td>
+
<td>0.2489</td>
+
<td>0.3852</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0141</td>
+
<td>0.0849</td>
+
<td>0.2246</td>
+
<td>0.4806</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device5</th>
+
<th>Colony 1</th>
+
<td>0.0118</td>
+
<td>0.1365</td>
+
<td>0.2730</td>
+
<td>0.4713</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0244</td>
+
<td>0.1875</td>
+
<td>0.3294</td>
+
<td>0.5378</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device6</th>
+
<th>Colony 1</th>
+
<td>0.0165</td>
+
<td>0.0906</td>
+
<td>0.2505</td>
+
<td>0.4839</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.0126</td>
+
<td>0.1120</td>
+
<td>0.2622</td>
+
<td>0.5442</td>
+
</tr>
+
</tbody>
+
</table>
+
<p><b>Table 6. Abs600 measurement after blank subtraction and correction.</b></p>
+
</figure>
+
<figure>
+
<div id="container-3"></div>
+
<p><b>Figure 3. Abs600 measurement after blank subtraction and correction.</b> We can find that two colony of device 1 are very different with other colonies.</p>
+
</figure>
+
<figure>
+
<table>
+
<thead>
+
<tr>
+
<th>Fluorescence</th>
+
<th>Replicate</th>
+
<th>0h</th>
+
<th>2h</th>
+
<th>4h</th>
+
<th>6h</th>
+
</tr>
+
</thead>
+
<tbody>
+
<tr>
+
<th rowspan="2">Negative control</th>
+
<th>Colony 1</th>
+
<td>8.4338</td>
+
<td>8.3597</td>
+
<td>8.3132</td>
+
<td>8.2557</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>8.1640</td>
+
<td>8.2490</td>
+
<td>8.2382</td>
+
<td>8.5850</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Positive Control</th>
+
<th>Colony 1</th>
+
<td>12.904</td>
+
<td>52.145</td>
+
<td>75.724</td>
+
<td>128.86</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>12.139</td>
+
<td>51.145</td>
+
<td>70.542</td>
+
<td>135.21</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device1</th>
+
<th>Colony 1</th>
+
<td>40.412</td>
+
<td>51.952</td>
+
<td>65.562</td>
+
<td>76.598</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>28.957</td>
+
<td>41.592</td>
+
<td>45.161</td>
+
<td>54.124</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device2</th>
+
<th>Colony 1</th>
+
<td>14.583</td>
+
<td>55.274</td>
+
<td>93.031</td>
+
<td>184.99</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>12.624</td>
+
<td>51.857</td>
+
<td>88.926</td>
+
<td>174.29</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device3</th>
+
<th>Colony 1</th>
+
<td>8.3959</td>
+
<td>8.6644</td>
+
<td>9.1428</td>
+
<td>9.9313</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>7.8957</td>
+
<td>8.2509</td>
+
<td>9.4031</td>
+
<td>11.753</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device4</th>
+
<th>Colony 1</th>
+
<td>10.542</td>
+
<td>55.101</td>
+
<td>81.432</td>
+
<td>118.27</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>10.275</td>
+
<td>50.620</td>
+
<td>76.936</td>
+
<td>135.14</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device5</th>
+
<th>Colony 1</th>
+
<td>8.1853</td>
+
<td>15.857</td>
+
<td>18.422</td>
+
<td>23.463</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>8.3564</td>
+
<td>18.187</td>
+
<td>20.175</td>
+
<td>26.058</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device6</th>
+
<th>Colony 1</th>
+
<td>7.9647</td>
+
<td>8.1582</td>
+
<td>8.2918</td>
+
<td>8.8606</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>7.8015</td>
+
<td>8.0578</td>
+
<td>8.2781</td>
+
<td>8.7673</td>
+
</tr>
+
</tbody>
+
</table>
+
<p><b>Table 7. Raw data of fluorescence measurement.</b></p>
+
</figure>
+
<figure>
+
<table>
+
<thead>
+
<tr>
+
<th>Fluorescence</th>
+
<th>Replicate</th>
+
<th>0h</th>
+
<th>2h</th>
+
<th>4h</th>
+
<th>6h</th>
+
</tr>
+
</thead>
+
<tbody>
+
<tr>
+
<th rowspan="2">Negative control</th>
+
<th>Colony 1</th>
+
<td>0.5088</td>
+
<td>0.4347</td>
+
<td>0.3882</td>
+
<td>0.3307</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.2390</td>
+
<td>0.3240</td>
+
<td>0.3132</td>
+
<td>0.6600</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Positive Control</th>
+
<th>Colony 1</th>
+
<td>4.9786</td>
+
<td>44.220</td>
+
<td>67.799</td>
+
<td>120.94</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>4.2144</td>
+
<td>43.220</td>
+
<td>62.617</td>
+
<td>127.29</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device1</th>
+
<th>Colony 1</th>
+
<td>32.487</td>
+
<td>44.027</td>
+
<td>57.637</td>
+
<td>68.673</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>21.032</td>
+
<td>33.667</td>
+
<td>37.236</td>
+
<td>46.199</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device2</th>
+
<th>Colony 1</th>
+
<td>6.6580</td>
+
<td>47.349</td>
+
<td>85.106</td>
+
<td>177.06</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>4.6994</td>
+
<td>43.932</td>
+
<td>81.001</td>
+
<td>166.37</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device3</th>
+
<th>Colony 1</th>
+
<td>0.4709</td>
+
<td>0.7394</td>
+
<td>1.2178</td>
+
<td>2.0063</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>-0.0293</td>
+
<td>0.3259</td>
+
<td>1.4781</td>
+
<td>3.8283</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device4</th>
+
<th>Colony 1</th>
+
<td>2.6174</td>
+
<td>47.176</td>
+
<td>73.507</td>
+
<td>110.34</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>2.3496</td>
+
<td>42.695</td>
+
<td>69.011</td>
+
<td>127.21</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device5</th>
+
<th>Colony 1</th>
+
<td>0.2603</td>
+
<td>7.9322</td>
+
<td>10.497</td>
+
<td>15.538</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>0.4314</td>
+
<td>10.262</td>
+
<td>12.250</td>
+
<td>18.133</td>
+
</tr>
+
<tr>
+
<th rowspan="2">Test Device6</th>
+
<th>Colony 1</th>
+
<td>0.0397</td>
+
<td>0.2332</td>
+
<td>0.3668</td>
+
<td>0.9356</td>
+
</tr>
+
<tr>
+
<th>Colony 2</th>
+
<td>-0.1235</td>
+
<td>0.1328</td>
+
<td>0.3531</td>
+
<td>0.8423</td>
+
</tr>
+
</tbody>
+
</table>
+
<p><b>Table 8. Fluorescence measurement after blank subtraction and correction.</b></p>
+
</figure>
+
<figure>
+
<div id="container-4"></div>
+
<p><b>Figure 4. Fluorescence measurement after blank subtraction and correction.</b></p>
+
</figure>
+
<figure>
+
<div id="container-5"></div>
+
<p>You can select area to zoom it.</p>
+
<p><b>Figure 5. Average level of devices.</b> In this figure, we find that the FI/Abs600 in device 1 is obviously higher than positive control,device 2,4 and 5. And another is that negative control and device 3,6 seems didn’t express GFP, or the expression is too low to observe.</p>
Many teachers and iGEM Headquarters says that reliable and repeatable measurement is a key component to all engineering disciplines. But to be honest it's difficult for laboratories all around the world to measure something in the same standard, so iGEM developing a robust measurement procedure for green fluorescent protein (GFP) every years to lead the team all over world could measure it.
It's the fourth year for iGEM to require teams which participate in finishing the interlab work. And GFP is a most widely used.
"All of the 2017 iGEM teams are invited and encouraged to participate in the Fourth International InterLaboratory Measurement Study in synthetic biology. We're hoping this study will get you excited for iGEM and help prepare you for the summer!" says by iGEM Headquarters. Actually, for us the interlab work not only to make fun but the more important part is that it can lead different laboratory to normalize their measure methods and machines so that we can use data from other lab easily, which will benefit our experiment and project a lot.
We all glad to participate in such significant international work with those friendly workmates all around world.
Materials and methods
Materials
Plasmid DNA (1ng in total, 100 pg/uL in 10uL of ddH20)
Positive control BBa_I20270
Negative control BBa_R0040
Test Device 1: J23101.BCD2.E0040.B0015
Test Device 2: J23106.BCD2.E0040.B0015
Test Device 3: J23117.BCD2.E0040.B0015
Test Device 4: J23101+I13504
Test Device 5: J23106+I13504
Test Device 6: J23117+I13504
Strain Used
Escherichia coli DH5α
Media Used
LB (Luria Bertani) media
Reagent Used
1xPBS (phosphate buffered saline)
Chloramphenicol (stock concentration 25 mg/mL dissolved in EtOH)
FITC Standard: one tube with dried down FITC for creating a FITC standard
LUDOX: one tube with 30% colloidal silica suspended in 1mL of water
Consumable Items
50 ml Falcon tube
1.5 ml eppendorf tubes for sample storage
Ice box with ice
Pipettes and tip
96 well plate
Machines
Thermo scientific
Varioskan Flash
CRYSTAL
Incubator Shaker
Yiheng-China
HZQ-F160A constant temperature incubator
AIRTECH
Vertical Flow Clean Bench
Methods
Calibration
OD600 Reference poin
FITC fluorescence standard curve
Cell measurement
Transformation
Measurements
Data and Analysis
Normalization work
LUDOX-HS40
H20
Replicate 1
0.0546438
0.47919
Replicate 2
0.0646164
0.039539
Replicate 3
0.0528094
0.062382
Replicate 4
0.0468351
0.054382
Arith.Mean
0.05472618
0.051055
Corrected Abs600
0.0036709
Reference OD600
0.00425
OD600/Abs600
11.5775423
Table 1. OD600 Reference Point.
Unit Scaling Factors:
OD600/Abs600
11.58
uM Fluorescein/a.u.
0.003458
Table 4. Normalization of initial concentration of colony. In order to unify the initial concentration before , we use Varioskan Flash to measure the OD600 after incubating overnight.
Cell Measurement
Because of the large account of data, we only use an average data in this form. (Average of 4 replicates of one colony)
Abs600
Replicate
0h
2h
4h
6h
Negative control
Colony 1
0.0666
0.1893
0.2708
0.4942
Colony 2
0.0615
0.1929
0.3000
0.5681
Positive Control
Colony 1
0.0601
0.1638
0.2949
0.5425
Colony 2
0.0613
0.1839
0.2982
0.5815
Test Device1
Colony 1
0.0647
0.0649
0.0757
0.0816
Colony 2
0.0560
0.0612
0.0627
0.0732
Test Device2
Colony 1
0.0616
0.1685
0.2480
0.4316
Colony 2
0.0607
0.1928
0.2977
0.4638
Test Device3
Colony 1
0.0463
0.1566
0.3120
0.4887
Colony 2
0.0521
0.1291
0.3464
0.6376
Test Device4
Colony 1
0.0571
0.1329
0.2899
0.4263
Colony 2
0.0551
0.1259
0.2656
0.5216
Test Device5
Colony 1
0.0528
0.1776
0.3140
0.5123
Colony 2
0.0655
0.2286
0.3704
0.5788
Test Device6
Colony 1
0.0575
0.1316
0.2915
0.5249
Colony 2
0.0536
0.1530
0.3032
0.5852
Tablet5. Raw Abs600 measurement.
Abs600
Replicate
0h
2h
4h
6h
Negative control
Colony 1
0.0256
0.1483
0.2298
0.4942
Colony 2
0.0205
0.1518
0.2590
0.5271
Positive Control
Colony 1
0.0191
0.1227
0.2539
0.5014
Colony 2
0.0203
0.1429
0.2571
0.5404
Test Device1
Colony 1
0.0237
0.0239
0.0347
0.0405
Colony 2
0.0149
0.0202
0.0217
0.0321
Test Device2
Colony 1
0.0206
0.1274
0.2069
0.3905
Colony 2
0.0197
0.1518
0.2567
0.4228
Test Device3
Colony 1
0.0053
0.1156
0.2710
0.4477
Colony 2
0.0110
0.0881
0.3053
0.5966
Test Device4
Colony 1
0.0161
0.0919
0.2489
0.3852
Colony 2
0.0141
0.0849
0.2246
0.4806
Test Device5
Colony 1
0.0118
0.1365
0.2730
0.4713
Colony 2
0.0244
0.1875
0.3294
0.5378
Test Device6
Colony 1
0.0165
0.0906
0.2505
0.4839
Colony 2
0.0126
0.1120
0.2622
0.5442
Table 6. Abs600 measurement after blank subtraction and correction.
Fluorescence
Replicate
0h
2h
4h
6h
Negative control
Colony 1
8.4338
8.3597
8.3132
8.2557
Colony 2
8.1640
8.2490
8.2382
8.5850
Positive Control
Colony 1
12.904
52.145
75.724
128.86
Colony 2
12.139
51.145
70.542
135.21
Test Device1
Colony 1
40.412
51.952
65.562
76.598
Colony 2
28.957
41.592
45.161
54.124
Test Device2
Colony 1
14.583
55.274
93.031
184.99
Colony 2
12.624
51.857
88.926
174.29
Test Device3
Colony 1
8.3959
8.6644
9.1428
9.9313
Colony 2
7.8957
8.2509
9.4031
11.753
Test Device4
Colony 1
10.542
55.101
81.432
118.27
Colony 2
10.275
50.620
76.936
135.14
Test Device5
Colony 1
8.1853
15.857
18.422
23.463
Colony 2
8.3564
18.187
20.175
26.058
Test Device6
Colony 1
7.9647
8.1582
8.2918
8.8606
Colony 2
7.8015
8.0578
8.2781
8.7673
Table 7. Raw data of fluorescence measurement.
Fluorescence
Replicate
0h
2h
4h
6h
Negative control
Colony 1
0.5088
0.4347
0.3882
0.3307
Colony 2
0.2390
0.3240
0.3132
0.6600
Positive Control
Colony 1
4.9786
44.220
67.799
120.94
Colony 2
4.2144
43.220
62.617
127.29
Test Device1
Colony 1
32.487
44.027
57.637
68.673
Colony 2
21.032
33.667
37.236
46.199
Test Device2
Colony 1
6.6580
47.349
85.106
177.06
Colony 2
4.6994
43.932
81.001
166.37
Test Device3
Colony 1
0.4709
0.7394
1.2178
2.0063
Colony 2
-0.0293
0.3259
1.4781
3.8283
Test Device4
Colony 1
2.6174
47.176
73.507
110.34
Colony 2
2.3496
42.695
69.011
127.21
Test Device5
Colony 1
0.2603
7.9322
10.497
15.538
Colony 2
0.4314
10.262
12.250
18.133
Test Device6
Colony 1
0.0397
0.2332
0.3668
0.9356
Colony 2
-0.1235
0.1328
0.3531
0.8423
Table 8. Fluorescence measurement after blank subtraction and correction.
Discussion
At first time, when we decide to transform plasmid into E.coli, what interesting is that we find device 1 can't growth in 170ug/ml or in 90ug/ml chloramphenicol at the same time we know exactly this plasmid is a high express one. Based on figure 3, we find that the growth curve of E.coli (except device 1) are all similar to the front part of S-Curve of Logistic regression, but it was strange that the two colony of device 1 seems don't change obviously in bacterial density which is different with other colonies. We conjectured this phenomenon came from the extremely high expression quantity of GFP which already produce severe cytotoxicity. As a result the growth of device 1 is very slow and can't growth in high concentration of chloramphenicol.
In figure 4 and figure 5, we analyzed the relationship between fluorescence and Abs 600. Then find that the promotor of GFP in device 1 actually is strongest in 8 plasmid which as 5 times as positive control, device 2 and device 4. The expression of GFP in device 5 seems very low, only one-tenth of positive control.
Also, we find that if the sampling time overlong, the E.coli growth will be restrain because of long time of low-temperature stress, in this suitcase, the growth of bacteria density is very slow and unparalleled in two colonies.