LOCUS       sequence-235305-        4279 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Functional test of ECF sigma factors and their cognate promoters.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1846 to 1880)
  AUTHORS   Huang HH, Qian Y, Del Vecchio D
  TITLE     A quasi-integral controller for adaptation of genetic modules to 
            variable ribosome demand.
  JOURNAL   Nat Commun. 2018 Dec 21;9(1):5415. doi: 10.1038/s41467-018-07899-z.
  PUBMED    30575748
REFERENCE   2  (bases 1 to 4279)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2018-12-21"; volume: "9(1):5415. doi"; pages: " 
            10.1038/s41467-018-07899-z"
COMMENT     Sequence Label: pHH03C_32
FEATURES             Location/Qualifiers
     source          1..4279
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..105
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(106..174,175..966)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLHKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(324..343)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      996..1090
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      1193..1738
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     1639..1658
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        1846..1880
                     /label=﻿BBa_J23100
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 1.0
                     "
                     /citation=[1]
     RBS             1896..1907
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1908..2621
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1938..1966)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2561..2581
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             2634..2645
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     promoter        2677..2711
                     /label=BBa_J23114
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.1"
     RBS             2720..2731
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2738..3361
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(2811..2830)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      3382..3468
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     protein_bind    3477..3495
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3502..3520
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             3539..3550
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             4133..4150
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4203..4221)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4217..4264
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
       61 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
      121 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
      181 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
      241 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
      301 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
      361 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
      421 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
      481 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
      541 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
      601 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
      661 acaacgttgc acaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
      721 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
      781 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
      841 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
      901 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
      961 tggtaactcg agctgatcct tcaagagctc gcttggactc ctgttgatag atccagtaat
     1021 gacctcagaa ctccatctgg atttgttcag aacgctcggt tgccgccggg cgttttttat
     1081 tggtgagaat ttacaactta tatcgtatgg ggctgacttc aggtgctaca tttgaagaga
     1141 taaattgcac tgaaatctag aaatatttta tctgattaat aagatgatct tcttgagatc
     1201 gttttggtct gcgcgtaatc tcttgctctg aaaacgaaaa aaccgccttg cagggcggtt
     1261 tttcgaaggt tctctgagct accaactctt tgaaccgagg taactggctt ggaggagcgc
     1321 agtcaccaaa acttgtcctt tcagtttagc cttaaccggc gcatgacttc aagactaact
     1381 cctctaaatc aattaccagt ggctgctgcc agtggtgctt ttgcatgtct ttccgggttg
     1441 gactcaagac gatagttacc ggataaggcg cagcggtcgg actgaacggg gggttcgtgc
     1501 atacagtcca gcttggagcg aactgcctac ccggaactga gtgtcaggcg tggaatgaga
     1561 caaacgcggc cataacagcg gaatgacacc ggtaaaccga aaggcaggaa caggagagcg
     1621 cacgagggag ccgccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     1681 ccactgattt gagcgtcaga tttcgtgatg cttgtcaggg gggcggagcc tatggaaaaa
     1741 cggctttgcc gcggccctct cacttccctg ttaagtatct tcctggcatc ttccaggaaa
     1801 tctccgcccc gttcgtaagc catttccgct cgccgcagtc gaacgttgac ggctagctca
     1861 gtcctaggta cagtgctagc ataacaaact ttgacaaaga ggagaaaatg cgtaaaggag
     1921 aagaactttt cactggagtt gtcccaattc ttgttgaatt agatggtgat gttaatgggc
     1981 acaaattttc tgtcagtgga gagggtgaag gtgatgcaac atacggaaaa cttaccctta
     2041 aatttatttg cactactgga aaactacctg ttccatggcc aacacttgtc actactttcg
     2101 gttatggtgt tcaatgcttt gcgagatacc cagatcatat gaaacagcat gactttttca
     2161 agagtgccat gcccgaaggt tatgtacagg aaagaactat atttttcaaa gatgacggga
     2221 actacaagac acgtgctgaa gtcaagtttg aaggtgatac ccttgttaat agaatcgagt
     2281 taaaaggtat tgattttaaa gaagatggaa acattcttgg acacaaattg gaatacaact
     2341 ataactcaca caatgtatac atcatggcag acaaacaaaa gaatggaatc aaagttaact
     2401 tcaaaattag acacaacatt gaagatggaa gcgttcaact agcagaccat tatcaacaaa
     2461 atactccaat tggcgatggc cctgtccttt taccagacaa ccattacctg tccacacaat
     2521 ctgccctttc gaaagatccc aacgaaaaga gagaccacat ggtccttctt gagtttgtaa
     2581 cagctgctgg gattacacat ggcatggatg aactatacaa ataataacca attattgaag
     2641 gccgctaacg cggccttttt ttgtttctgg tctcccttta tggctagctc agtcctaggt
     2701 acaatgctag caattcatta aagaggagaa aggtaccatg tccagattag ataaaagtaa
     2761 agtgattaac agcgcattag agctgcttaa tgaggtcgga atcgaaggtt taacaacccg
     2821 taaactcgcc cagaagctag gtgtagagca gcctacattg tattggcatg taaaaaataa
     2881 gcgggctttg ctcgacgcct tagccattga gatgttagat aggcaccata ctcacttttg
     2941 ccctttagaa ggggaaagct ggcaagattt tttacgtaat aacgctaaaa gttttagatg
     3001 tgctttacta agtcatcgcg atggagcaaa agtacattta ggtacacggc ctacagaaaa
     3061 acagtatgaa actctcgaaa atcaattagc ctttttatgc caacaaggtt tttcactaga
     3121 gaatgcatta tatgcactca gcgctgtggg gcattttact ttaggttgcg tattggaaga
     3181 tcaagagcat caagtcgcta aagaagaaag ggaaacacct actactgata gtatgccgcc
     3241 attattacga caagctatcg aattatttga tcaccaaggt gcagagccag ccttcttatt
     3301 cggccttgaa ttgatcatat gcggattaga aaaacaactt aaatgtgaaa gtgggtccta
     3361 ataaacgcgt gctagaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt
     3421 cgttttatct gttgtttgtc ggtgaacgct ctcctgagta ggacaaatta ctagagtccc
     3481 tatcagtgat agagattgac atccctatca gtgatagaga tactgagcac tactagagaa
     3541 agaggagaaa tactagatga ccgaaattca tctgcagacc accgaaagca ccagcgttaa
     3601 tgatggtctg ccgctgaata ttgattggga aggtattttt cgtgaacatg gtcgtcgtgt
     3661 gcatcatttt attcgtaaac gtgttagcca tcgtgaagat gccgaagatc tggaacaaat
     3721 gacctggctg gaagttctgc gtaatcgtga taaatttgcc ggtgcaagcc gtccggaaac
     3781 ctgggttttt ggtattgcac tgaatctggt gcgtaatcat tttcgtctgc agagcggtcg
     3841 tccgcgttgt gatgaactgg aagatgatat tattctgacc cagggtgatg atccgagcca
     3901 tattaccgaa tatcagcgtc tgctgaatag caccctgaat agtattgcaa atctgccgga
     3961 agatacccgt cgcctgctga acatgctggt tgaaaaagat ggtagctatc aggcaattgc
     4021 agcccatctg aatattccga ttggcaccgt tcgtagccgt ctgagccgtg cacgtgttac
     4081 cctgaaacag agcgttttta gctaaggatc caagcttgcg gccgcactcg agcaccacca
     4141 ccaccaccac tgagatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc
     4201 caccgctgag caataactag cataacccct tggggcctct aaacgggtct tgaggggttt
     4261 tttgggcgtc tccactgtg
//
