Part:BBa_K3192008

Custom RBS Optimzed for BBa_K2259018

Synthetic Ribosomal Binding site for BBa_K2259018, a BioBrick created for split kanamycin resistance to maintain two plasmids within one cell using a single antibiotic resistance gene.

The 2019 Virginia iGEM Team sought efficiency and productivity in designing their genetic construct as the device required two plasmids to operate. Two plasmids bears a large metabolic load on the chassis¹, so, synthetic ribosomal binding sites were developed in order to optimize translations rates for the genes used in the styrene-degrading and polyhydroxybutyrate-producing plasmids. This ribosomal binding site is optimized for part BBa_K2259018 with a translation initiation rate of 306146.74.

When expressing gene clusters in a foreign chassis, the endogenous ribosomal binding sites (RBS) are often included in the sequence information provided by NCBI. Although these RBS are sufficient, they may not provide optimal translational efficiency. Synthetic RBSs provide a solution to increase translation initiation rates by using software to design a RBS with a linear secondary structure that allows for optimal ribosomal binding and translation. Our team’s project required enhanced translation of our coding sequences; therefore we chose to optimize all RBSs for high translation initiation rates. Using DeNovo DNA, our team created synthetic RBSs that ran thousands of iterations to create substantially elevated translation initiation rates.

References:

1) Watve, M. M., Dahanukar, N. & Watve, M. G. Sociobiological control of plasmid copy number in bacteria. PloS one (2010). Available at: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2827543/. (Accessed: 22nd October 2019)

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]