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Part by part: synthetic biology parts used in solventogenic Clostridia Ivan Slavkov Gyulev, Benjamin J Willson, Rosanna Catherine Hennessy, Preben Krabben, Elizabeth R Jenkinson, and Gavin Hugh Thomas ACS Synth. Biol., Just Accepted Manuscript • DOI: 10.1021/acssynbio.7b00327 • Publication Date (Web): 29 Nov 2017 Downloaded from http://pubs.acs.org on December 1, 2017

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ACS Synthetic Biology is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

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Page 55 of 57 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 1. Phylogenetic tree of 16S sequences from selected organisms from the genus Clostridium. The tree was built using Maximum Likelihood method based on the Tamura-Nei model. The bootstrap consensus tree inferred from 1000 replicates is taken to represent the evolutionary history of the taxa analyzed. Evolutionary analyses were conducted in MEGA5. Green circles denote butanol-producing species used in industrial biotechnology (IB), red squares mark risk group 2 species (risk group 3 being highest risk). 142x210mm (96 x 96 DPI)



Figure 3. Application of terminator prediction software to C. acetobutylicum. A. Outputs from predicted terminator sets derived from the C. acetobutylicum genome by application of TransTermHP (TTHP), RNIE and WedGeSTer were compared. B. An alignment of predictions of a putative bi-directional terminator (marked by T sign) by all three algorithms (note all three produce separate predictions for plus (+)and minus (-))C. RNA structures predicted by RNAfold for the TransTermHP sequence, drawn with VARNA; it is worth noting that the base-pairing between the A-tract and U-tract may not form until after termination. Terminator lengths: WG: maximum length (max)-48bp, minimum (min) - 28bp, average (avg) - 35.8bp; RNIE: max-45bp, min- 34bp, avg-41.7bp,TTHP: max-89bp,min-41bp,avg-57.2bp. 254x190mm (96 x 96 DPI)


Page 56 of 57

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