Naming of standards of physical composition of BioBrick parts

This BioBricks Foundation Request for Comments (BBF RFC) proposes a naming convention for standards of physical composition of BioBrick parts in which assembly standards are named according to the BBF RFC that specifies the standard

Instructions to BBF RFC Authors

This BioBricks Foundation Request for Comments (BBF RFC) provides in- formation about the preparation and submission of BBF RFCs to The Bio- Bricks Foundation (BBF)

Comparative analysis of three studies measuring fluorescence from engineered bacterial genetic constructs

Reproducibility is a key challenge of synthetic biology, but the foundation of reproducibility is only as solid as the reference materials it is built upon. Here we focus on the reproducibility of fluorescence measurements from bacteria transformed with engineered genetic constructs. This comparative analysis comprises three large interlaboratory studies using flow cytometry and plate readers, identical genetic constructs, and compatible unit calibration protocols. Across all three studies, we find similarly high precision in the calibrants used for plate readers. We also find that fluorescence measurements agree closely across the flow cytometry results and two years of plate reader results, with an average standard deviation of 1.52-fold, while the third year of plate reader results are consistently shifted by more than an order of magnitude, with an average shift of 28.9-fold. Analyzing possible sources of error indicates this shift is due to incorrect preparation of the fluorescein calibrant. These findings suggest that measuring fluorescence from engineered constructs is highly reproducible, but also that there is a critical need for access to quality controlled fluorescent calibrants for plate readers

Reproducibility of fluorescent expression from engineered biological constructs in E. coli

We present results of the first large-scale interlaboratory study carried out in synthetic biology, as part of the 2014 and 2015 International Genetically Engineered Machine (iGEM) competitions. Participants at 88 institutions around the world measured fluorescence from three engineered constitutive constructs in E. coli. Few participants were able to measure absolute fluorescence, so data was analyzed in terms of ratios. Precision was strongly related to fluorescent strength, ranging from 1.54-fold standard deviation for the ratio between strong promoters to 5.75-fold for the ratio between the strongest and weakest promoter, and while host strain did not affect expression ratios, choice of instrument did. This result shows that high quantitative precision and reproducibility of results is possible, while at the same time indicating areas needing improved laboratory practices

Correction: Reproducibility of Fluorescent Expression from Engineered Biological Constructs in E. coli.

[This corrects the article DOI: 10.1371/journal.pone.0150182.]

Constitutive fluorescence constructs measured in the 2014 and 2015 iGEM Interlab Studies.

<p>Constitutive fluorescence constructs measured in the 2014 and 2015 iGEM Interlab Studies.</p

Instrument-to-instrument variation within a single team.

<p>Each column represents a set of replicates measured by the same laboratory on different instruments. Blue circles are the ratios for individual instruments, normalized by mean ratio; the red line spans ±1 std.dev.</p

Relationship between mean ratio and precision.

<p>The lower another promoter’s expression is in relation to the strong construct, the higher the variation in measurement: standard deviation grows approximately in proportion to the square root of the mean ratio.</p

Strains reported in 2014 and 2015 iGEM Interlab Studies.

<p>Strains reported in 2014 and 2015 iGEM Interlab Studies.</p

Distributions of data partitioned by <i>E. coli</i> strain for each ratio, omitting outliers.

<p>Note that the BL21 subset in 2014 has only a single non-outlier data point and may thus be effectively ignored.</p