Coding

Part:BBa_K4268000:Design

Designed by: J. Aubrey, J. Alvarenga, L. Buchanan, J. Reyes, D. Yashinski   Group: iGEM22_SUNY_Oneonta   (2022-07-15)
Revision as of 23:02, 25 September 2022 by Jazmine124 (Talk | contribs) (→‎Design Notes)


S-TIP37 Head-Tail Connector


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 346
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

Synechococcus sp WH 8109, was decided as the desired cyanobacterial species that SUNY_Oneonta iGEM would like to target, S-TIP 37 is the infecting cyanophage of this cyanobacterial species. Although S-TIP 37's genome has been fully sequenced there is little experimental evidence of research on the genome, thus resulting in many hypothetical proteins whose functions are unknown. To better characterize these putative genes, a series of bioinformatic tools were employed.


SUNY_Oneonta iGEM identified another T7-like phage that has had considerable research done with its genome, Syn-5. The structural and genomic similarities between the Syn-5 and S-TIP 37, allowed direct comparisons to be drawn between the two. Utilizing NCBI's BLAST tool SUNY_Oneonta iGEM was able to genetically compare the suspected Head-Tail Connector putative gene of S-TIP 37 with that of Syn-5's confirmed one. There was significant positive alignment between the two genomes, allowing SUNY_Oneonta to conclude that the S-TIP 37 gene is likely the Head-Tail Connector.


Insert poster table comparison Caption: A BLAST alignment comparing S-TIP 37 and Syn-5's Head-Tail Connector genes. Similarities are indicated in red and differences in blue. BLAST has determined a 88% similarity in identities between the two.

Upon the isolation of the Head-Tail Connector sequence, the following edits were made:

  • The sequence was downloaded and underwent codon optimization to ensure RCF10 and RCF1000 compatibility. The following restriction sites were domesticated:
    • XbaI at nucleotide positions 54-59
    • SapI at nucleotide positions 205-211
    • SapI at nucleotide positions 265-272
  • To complete Level 0 Golden Gate Assembly, the sequence contains SapI sites at the ends

Source

The source of this sequence is from the S-TIP37 genome (GenBanK: NC_048026.1)

References

The S-TIP37 genome was found on GenomeNet