Difference between revisions of "Part:BBa K1323007"

Revision as of 02:20, 17 October 2014

sRNA (3): YFP-targeting molecule

Small RNAs (sRNAs) are small double-stranded transcripts capable of silencing mRNAs of complementary sequence (Yoo et al., 2013). The sRNA sequence was designed with two functional parts: the part that is complementary to the target mRNA (target binding site) and MicC scaffold region for recruitment of Hfq BBa_K1323019), a protein which enhances the sRNA to target mRNA hybridization (Yoo et al., 2013). The MicC RNA sequence is the best scaffold sequence as shown by previous studies done by the same group (Na et al., 2013). Since the target mRNA is YFP, the target binding sequence of this sRNA is complementary to the S. epidermidis codon optimized YFP BBa_K1323010.

sRNA 3 targets 28 bps downstream from the start codon of the YFP construct. This sequence has binding energy of -30.3 kcal/mol as determined using DINAMelt software (http://mfold.rna.albany.edu/?q=DINAMelt/Two-state-melting). For effective binding, sRNA should have binding energy between -30kcal/mol to -40kcal/mol (Yoo et al., 2013). We aligned sRNA 3 sequence with other constructs and ATCC 12228 genome to determine the highest binding energy for non-specific RNA binding, which is -13.03 kcal/mol. This shows that sRNA1 binds more specifically to YFP and should not interfere with other gene expressions in our system.

This part was DNA-Synthesized

References

Dokyun Na, S. M. (2013). Metabolic engineering of Escherichia coli using synthetic small regulatory RNAs. Nature Biotechnology, 170-174.

Na, D. et al (2013). Metabolic engineering of Escherichia coli using synthetic small regulatory RNAs. Nature Biotechnology 31, 170–174.

Yoo, S, M., Na, D., Lee, S. Y. (2013). Design and use of synthetic regulatory small RNAs to control gene expression in Escherichia coli. Nature protocol 8(9), 1694-1707.

Sequence and Features