Regulatory

Part:BBa_K1484341:Design

Designed by: Angelina Munabi   Group: iGEM14_Cambridge-JIC   (2014-10-10)


P_NiI9, marchantia promoter


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 155
    Illegal EcoRI site found at 861
    Illegal EcoRI site found at 1001
    Illegal XbaI site found at 758
    Illegal PstI site found at 1752
    Illegal PstI site found at 1763
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 155
    Illegal EcoRI site found at 861
    Illegal EcoRI site found at 1001
    Illegal PstI site found at 1752
    Illegal PstI site found at 1763
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 155
    Illegal EcoRI site found at 861
    Illegal EcoRI site found at 1001
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 155
    Illegal EcoRI site found at 861
    Illegal EcoRI site found at 1001
    Illegal XbaI site found at 758
    Illegal PstI site found at 1752
    Illegal PstI site found at 1763
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 155
    Illegal EcoRI site found at 861
    Illegal EcoRI site found at 1001
    Illegal XbaI site found at 758
    Illegal PstI site found at 1752
    Illegal PstI site found at 1763
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

The removal of illegal restriction sites was considered but not completed due to direct interaction of the DNA with regulatory proteins. Also, it was ensured that the region selected for this part was directly upstream of the first ATG of the gene used to identify the promoter sequence. This was so that the 5' UTR, that is vital in plants, was maintained.


Source

This part was found by screening the genome of the Marchantia polymorpha Cam strain maintained by the Haseloff lab for homologues to Nitrate Transporter proteins in A. Thaliana and B. Napus, known respectively as AtNRT2.1 or ACH1 and BnNRT1.2 or RCH2. Transcription is thought to be induced by nitrate (NO-3). Matches to the protein CDS sequence were found using Geneious to perform a tblastn search on the genome scaffolds. Predicted genes that contained hits graded above 30% and with at least 40% congruence to mRNA transcript sequences were shortlisted. The best gene candidates (judged according to number and distribution of hits along its length, and supporting mRNA sequence) formed the basis for our predicted promoters. This part was isolated from a 2kb region upstream of the first ATG of such a gene.


References

Forde BG. 2000. Nitrate transporters in plants: structure, function and regulation. (BBA) – Biomembranes 1465(1-2):219-235. Filleur S, Daniel-Vedele F. 1998. Expression analysis of a high-affinity nitrate transporter isolated from Arabidopsis thaliana by differential display. Planta 207: 461-469. Thalemine, ATG id AT1G08090 GenBank, NIH. Accession no: U17987