Coding

Part:BBa_M12066:Design

Designed by: Abhinav Jain   Group: MIT BE 20.20 - S09 MIT BE 20.20 - S10   (2009-05-12)

Cellulose Synthase Gene


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
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 322
    Illegal SapI site found at 708


Design Notes

In order to select the sequence to use for the part, NCBI's ORF finder web application was used to determine the correct open reading frame to use. This yielded the sequence that will be used for the part. There were modifications that needed to be made in order to fix restriction enzyme capability. A CTG leucine codon occurring in a PST1 site at bp 325 was modified to CTC. This allowed RFC(10) and RFC(23) compatibility. BgIII site occurred at bp 852, however due to presence of Tryptophan (single codon) at 926, BamHI site could not be removed, and RFC(21) remains incompatible. GAA codon for Glu changed to GAG. At 1534 acc codon was changed to act for threonine. This allowed RFC(25) compatibility.There was a BamHI site at bp 926. Since the amino acid encoded at that position was tryptophan, we could not modify the tryptophan codon, so the flanking isoleucine codons were modified from atc to ata, removing the bamHI site.

Source

This part comes from the GenBank database, Accession number EU889445. It was found using a partial C. Reinhardtii sequence, and then utilizing NCBI's BLAST 2 tool to identify the closest matching complete gene sequence. This sequence was an E. Coli cellulose synthase, and it is this gene that has been used in creating this part. This part should be usable across species, as it has very close BLAST matches to genes from an assortment of cellulose producing microoragnisms, including green algae and various bacterial strains.

References