Difference between revisions of "Part:BBa P1014:Design"

(→‎Design Notes)
(→‎Design Notes)
 
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*XbaI site at 660 removed by mutating c->t which also improves codon usage.
 
*XbaI site at 660 removed by mutating c->t which also improves codon usage.
 
**gatgatttttacagggatattttagagaacgaaatctac,P1014-XF
 
**gatgatttttacagggatattttagagaacgaaatctac,P1014-XF
−
**gatgatttttacagggatattttagagaacgaaatctactt,P1014-XF2
+
**gatgatttttacagggatattttagagaacgaaatctact,P1014-XF2
 
**gtagatttcgttctctaaaatatccctgtaaaaatcatc,P1014-XR
 
**gtagatttcgttctctaaaatatccctgtaaaaatcatc,P1014-XR
  

Latest revision as of 03:23, 25 July 2006


gentamicin resistance cassette aph(2)


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]


Design Notes

  • PCR primers
    • gtttcttcgaattcgcggccgcttctagag gagataggttatgcaagat,P1014-F
    • gtttcttcctgcagcggccgctactagta gattccggattctaaaaaaagg,P1014-R2 (after terminator) [Bad, extra A]
    • gtttcttcctgcagcggccgctactagta ctatttaattttaatgcttctg,P1014-R3 (after stop codon)
    • gtttcttcctgcagcggccgctactagta gattccggattctaaaaaaggattgcta,P1014-R4
    • gtttcttcctgcagcggccgctactagta gattccggattctaaaaaagg,P1014-R5


  • XbaI site at 660 removed by mutating c->t which also improves codon usage.
    • gatgatttttacagggatattttagagaacgaaatctac,P1014-XF
    • gatgatttttacagggatattttagagaacgaaatctact,P1014-XF2
    • gtagatttcgttctctaaaatatccctgtaaaaatcatc,P1014-XR

Source

  • Novel gentamicin resistance gene
  • Acquired by PCR from pTEX5500ts, Genbank DQ208936, kind gift of Barbara E. Murray.
  • pTEX5500ts was, in turn was derived from Genbank AF016483.

References

  • Nallapareddy SR, Singh KV, Murray BE. Construction of improved temperature-sensitive and mobilizable vectors and their use for constructing mutations in the adhesin-encoding acm gene of poorly transformable clinical Enterococcus. Appl Environ Microbiol. 2006 Jan;72(1):334-45 faecium strains. PMID 16391062
  • Tsai SF, Zervos MJ, Clewell DB, Donabedian SM, Sahm DF, Chow JW. A new high-level gentamicin resistance gene, aph(2)-Id, in Enterococcus spp. Antimicrob Agents Chemother. 1998 May;42(5):1229-32. PMID 9593155