LOCUS       sequence-311388-        3737 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression (sigma 70 promoter) of GCaMP6f fluorophore 
            in E. coli..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 69 to 103)
  AUTHORS   Bruni GN, Kralj JM
  TITLE     Membrane voltage dysregulation driven by metabolic dysfunction 
            underlies bactericidal activity of aminoglycosides.
  JOURNAL   eLife. 2020 Aug 4;9. pii: 58706. doi: 10.7554/eLife.58706.
  PUBMED    32748785
REFERENCE   2  (bases 1 to 3737)
  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: "eLife."; 
            date: "2020-08-4"; volume: "9. pii: 58706. doi"; pages: " 
            10.7554/eLife.58706"
COMMENT     Sequence Label: pKL11-GCaMP6f bb100
FEATURES             Location/Qualifiers
     source          1..3737
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(1..44)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    47..68
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     promoter        69..103
                     /label=﻿BBa_J23100
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 1.0
                     "
                     /citation=[1]
     CDS             join(134..142,143..160,161..163,164..196,197..199,200..223,
                     224..250,251..307,308..313,314..583,584..601,602..1036,
                     1037..1042,1043..1486)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEEFSLFNKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      230..239
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     520..541
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(647..668)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(908..927)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     complement(1539..1556)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     primer_bind     complement(1753..1770)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1924..2512)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2004..2023)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2683..3474,3475..3543))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3306..3325
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3544..3648)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3716..3734
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 atttcagata aaaaaaatcc ttagctttcg ctaaggatga tttctggaat tcgcggccgc
       61 ttctagagtt gacggctagc tcagtcctag gtacagtgct agcattaact ttaagaagga
      121 gatatacata cccatgggtt ctcatcatca tcatcatcat ggtatggcta gcatgactgg
      181 tggacagcaa atgggtcggg atctgtacga cgatgacgat aaggatctcg ccaccatggt
      241 cgactcatca cgtcgtaagt ggaataagac aggtcacgca gtcagagcta taggtcggct
      301 gagctcactc gagaacgtct atatcaaggc cgacaagcag aagaacggca tcaaggcgaa
      361 cttcaagatc cgccacaaca tcgaggacgg cggcgtgcag ctcgcctacc actaccagca
      421 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcgtgca
      481 gtccaaactt tcgaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
      541 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagggcggta ccggagggag
      601 catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
      661 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggtgagggcg atgccaccta
      721 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
      781 cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa
      841 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac atccaggagc gcaccatctt
      901 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
      961 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
     1021 caagctggag tacaacctgc cggaccaact gactgaagag cagatcgcag aatttaaaga
     1081 ggaattctcc ctatttaaca aggacgggga tgggacaata acaaccaagg agctggggac
     1141 ggtgatgcgg tctctggggc agaaccccac agaagcagag ctgcaggaca tgatcaatga
     1201 agtagatgcc gacggtgacg gcacaatcga cttccctgag ttcctgacaa tgatggcaag
     1261 aaaaatgaaa tacagggaca cggaagaaga aattagagaa gcgttcggtg tgtttgataa
     1321 ggatggcaat ggctacatca gtgcagcaga gcttcgccac gtgatgacaa accttggaga
     1381 gaagttaaca gatgaagagg ttgatgaaat gatcagggaa gcagacatcg atggggatgg
     1441 tcaggtaaac tacgaagagt ttgtacaaat gatgacagcg aagtaagttc aaacggtctc
     1501 cagcttggct gttttggcgg atgagagaag attttcagcc tgatacagat taaatcagaa
     1561 cgcagaagcg gtctgataaa acagaatttg cctggcggca gtagcgcggt ggtcccacct
     1621 gaccccatgc cgaactcaga agtgaaacgc cgtagcgccg atggtagtgt ggggtctccc
     1681 catgcgagag tagggaactg ccaggcatca aataaaacga aaggctcagt cgaaagactg
     1741 ggccttggct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     1801 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     1861 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     1921 gtttttccac aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     1981 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     2041 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     2101 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     2161 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     2221 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     2281 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     2341 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     2401 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     2461 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     2521 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     2581 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     2641 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     2701 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     2761 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     2821 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     2881 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     2941 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     3001 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     3061 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     3121 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     3181 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     3241 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     3301 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     3361 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     3421 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     3481 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     3541 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     3601 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     3661 aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag
     3721 gcgtatcacg aggcaga
//
