Difference between revisions of "Team:Peking/InterLab"
| Line 33: | Line 33: | ||
margin-bottom: 0px; | margin-bottom: 0px; | ||
} | } | ||
| + | |||
| + | ol{ | ||
| + | list-style: none | ||
| + | } | ||
| + | |||
| + | ol li:before { | ||
| + | content: counter(ol) "."; | ||
| + | counter-increment: ol; | ||
| + | color: #e06e73; | ||
| + | text-align: right; | ||
| + | display: inline-block; | ||
| + | min-width: 1em; | ||
| + | margin-right: 0.5em | ||
| + | } | ||
| + | |||
| + | ol ol { | ||
| + | margin-left: 1.25714em | ||
| + | } | ||
| + | |||
| + | ol li { | ||
| + | text-indent: -1.55em; | ||
| + | line-height : 2em; | ||
| + | } | ||
| + | |||
| + | |||
| + | |||
| + | |||
| + | <style> | ||
| + | |||
| + | |||
| + | </style> | ||
| + | |||
</style> | </style> | ||
| Line 151: | Line 183: | ||
style="background: url('https://static.igem.org/mediawiki/2017/5/52/Peking_Interlab_background2.jpeg') center / cover; padding-left: 50px;"> | style="background: url('https://static.igem.org/mediawiki/2017/5/52/Peking_Interlab_background2.jpeg') center / cover; padding-left: 50px;"> | ||
<div class="mdl-card__title-text"> | <div class="mdl-card__title-text"> | ||
| − | <h1 style="font-size: xx-large; color: white"><strong>Interlab</strong></h1> | + | <h1 style="font-size: xx-large; color: white;text-shadow:2px 2px 8px #070707;"><strong>Interlab</strong></h1> |
</div> | </div> | ||
</div> | </div> | ||
| Line 173: | Line 205: | ||
</nav> | </nav> | ||
</section> | </section> | ||
| + | |||
| + | |||
<br> | <br> | ||
The Peking iGEM 2017 team is participating in the Fourth InterLaboratory Measurement Study in synthetic | The Peking iGEM 2017 team is participating in the Fourth InterLaboratory Measurement Study in synthetic | ||
| Line 186: | Line 220: | ||
<h2 id = "p2">Materials</h2> | <h2 id = "p2">Materials</h2> | ||
<h3>Plasmids</h3> | <h3>Plasmids</h3> | ||
| − | <ol> | + | <ol start="1"> |
| + | <script type="text/javascript"> | ||
| + | (function () { | ||
| + | |||
| + | var doc_ols = document.getElementsByTagName("ol"); | ||
| + | |||
| + | for (i = 0; i < doc_ols.length; i++) { | ||
| + | |||
| + | var ol_start = doc_ols[i].getAttribute("start") - 1; | ||
| + | doc_ols[i].setAttribute("style", "counter-reset:ol " + ol_start + ";"); | ||
| + | |||
| + | } | ||
| + | |||
| + | })(); | ||
| + | </script> | ||
<li>Positive Control (BBa_I20270): well 21B</li> | <li>Positive Control (BBa_I20270): well 21B</li> | ||
<li>Negative Control (BBa_R0040): well 21D</li> | <li>Negative Control (BBa_R0040): well 21D</li> | ||
| Line 200: | Line 248: | ||
<h3>Reagent</h3> | <h3>Reagent</h3> | ||
<ul type=circle> | <ul type=circle> | ||
| + | |||
| + | |||
<li>1ml LUDOX</li> | <li>1ml LUDOX</li> | ||
<li>ddH<SUB>2</SUB>O</li> | <li>ddH<SUB>2</SUB>O</li> | ||
| Line 208: | Line 258: | ||
<h3>Instruments</h3> | <h3>Instruments</h3> | ||
<ul type=circle> | <ul type=circle> | ||
| + | |||
| + | |||
<li>Pipettes</li> | <li>Pipettes</li> | ||
<li>96-well plate</li> | <li>96-well plate</li> | ||
| Line 213: | Line 265: | ||
<li>1.5-ml Eppendorf tubes for sample storage, ice bucket with ice</li> | <li>1.5-ml Eppendorf tubes for sample storage, ice bucket with ice</li> | ||
</ul> | </ul> | ||
| − | + | <br> | |
<h2 id = "p3">Methods</h2> | <h2 id = "p3">Methods</h2> | ||
At first, we used LUDOX-S40 as a single-point reference to obtain a ratiometric conversion factor to | At first, we used LUDOX-S40 as a single-point reference to obtain a ratiometric conversion factor to | ||
Revision as of 06:44, 29 October 2017
Interlab
The Peking iGEM 2017 team is participating in the Fourth InterLaboratory Measurement Study in synthetic biology, along with teams from around the world. We introduced eight plasmids into E. coli K-12 DH5-alpha and measured the GFP expression levels using a plate reader.
Introduction
Reliable and repeatable measurement is a key component of synthetic biology. However, there have been few opportunities to repeat the same measurements in different labs. In order to quantify the degree of variability exhibited by engineered genetic constructs across different laboratories, the measurement committee invited all iGEM teams to participate in the Interlab study, which provides researchers with a detailed protocol and data analysis form, with the aim to produce common, comparable units for measuring GFP on different plate readers.Materials
Plasmids
- Positive Control (BBa_I20270): well 21B
- Negative Control (BBa_R0040): well 21D
- Test Device 1 (BBa_J364000): well 21F
- Test Device 2 (BBa_J364001): well 21H
- Test Device 3 (BBa_J364002): well 21J
- Test Device 4 (BBa_J364003): well 21L
- Test Device 5 (BBa_J364004): well 21N
- Test Device 6 (BBa_J364005): well 21P
Strains
Escherichia coli strain DH5αReagent
- 1ml LUDOX
- ddH2O
- Fluorescein
- 10ml 1xPBS
Instruments
- Pipettes
- 96-well plate
- 50 ml Falcon tube (or equivalent, preferably amber or covered in foil to block light)
- 1.5-ml Eppendorf tubes for sample storage, ice bucket with ice
Methods
At first, we used LUDOX-S40 as a single-point reference to obtain a ratiometric conversion factor to transform our absorbance readings into standard OD600 data. Simultaneously, the ratiometric conversion factor accounts for instrument differences.Secondly, we generated a standard curve of fluorescence as a function of fluorescein concentration. We can thus use the standard curve to convert the fluorescence of GFP into a concentration of GFP.
Last and most importantly, we measured the A600 and fluorescence of green fluorescent protein. The GFP is used as a measurement marker of promoter activity, and the fluorescence/OD600 ratio is used to give an adjustment of the relative expression per cell. Furthermore, we tested some RBS devices that are intended to make gene expression more precise and reliable.
Results
Samples were prepared according to the standard protocol.The results for E. coli DH5α are shown in figures 4-8.Back < Lab Goto > Lab/...
