LOCUS       sequence-457220-        7041 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Sensor cassette FIRE2, sensor in backwards orientation, FIRE 
            fluorescent ratiometric sensor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7041)
  AUTHORS   Sangokoya C
  TITLE     The FIRE biosensor illuminates iron regulatory protein activity and 
            cellular iron homeostasis.
  JOURNAL   Cell Rep Methods. 2025 Jan 11:100960. doi: 
            10.1016/j.crmeth.2024.100960.
  PUBMED    39824193
REFERENCE   2  (bases 1 to 7041)
  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: "Cell Rep 
            Methods. 2025 Jan 11:100960. doi: 10.1016/j.crmeth.2024.100960."
COMMENT     Sequence Label: FIRESFLIP-PBCS-E670
FEATURES             Location/Qualifiers
     source          1..7041
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        228..1406
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          459..1397
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1354..1374
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1438..1447
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3161..3385
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3758..3792
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        complement(3941..3959)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3980..3996)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3980..3996)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3993..4015)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4004..4020
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4028..4034,4035..4052,4053..4058))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4073..4094
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4211..4228)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4382..4970)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4462..4481)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5141..5932,5933..6001))
                     /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     5764..5783
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6002..6106)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6132..6587)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6269..6290)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6481..6500
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6714..6736
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6728..6745
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6729..6745
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6752..6771
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6752..6770
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     repeat_region   6837..6899
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
ORIGIN
        1 caaaatttct tctataaagt aacaaaactt ttaaacattc tctcttttac aaaaataaac
       61 ttattttgta ctttaaaaac agtcatgttg tattataaaa taagtaatta gcttaaccta
      121 tacataatag aaacaaatta tacttattag tcagtcagaa acaactttgg cacatatcaa
      181 tattatgctc tcgttaatta agaattcgct agcccctgta cacgtgaggc tccggtgccc
      241 gtcagtgggc agagcgcaca tcgcccacag tccccgagaa gttgggggga ggggtcggca
      301 attgaaccgg tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat gtcgtgtact
      361 ggctccgcct ttttcccgag ggtgggggag aaccgtatat aagtgcagta gtcgccgtga
      421 acgttctttt tcgcaacggg tttgccgcca gaacacaggt aagtgccgtg tgtggttccc
      481 gcgggcctgg cctctttacg ggttatggcc cttgcgtgcc ttgaattact tccacctggc
      541 tgcagtacgt gattcttgat cccgagcttc gggttggaag tgggtgggag agttcgaggc
      601 cttgcgctta aggagcccct tcgcctcgtg cttgagttga ggcctggcct gggcgctggg
      661 gccgccgcgt gcgaatctgg tggcaccttc gcgcctgtct cgctgctttc gataagtctc
      721 tagccattta aaatttttga tgacctgctg cgacgctttt tttctggcaa gatagtcttg
      781 taaatgcggg ccaagatctg cacactggta tttcggtttt tggggccgcg ggcggcgacg
      841 gggcccgtgc gtcccagcgc acatgttcgg cgaggcgggg cctgcgagcg cggccaccga
      901 gaatcggacg ggggtagtct caagctggcc ggcctgctct ggtgcctggc ctcgcgccgc
      961 cgtgtatcgc cccgccctgg gcggcaaggc tggcccggtc ggcaccagtt gcgtgagcgg
     1021 aaagatggcc gcttcccggc cctgctgcag ggagctcaaa atggaggacg cggcgctcgg
     1081 gagagcgggc gggtgagtca cccacacaaa ggaaaagggc ctttccgtcc tcagccgtcg
     1141 cttcatgtga ctccacggag taccgggcgc cgtccaggca cctcgattag ttctcgagct
     1201 tttggagtac gtcgtcttta ggttgggggg aggggtttta tgcgatggag tttccccaca
     1261 ctgagtgggt ggagactgaa gttaggccag cttggcactt gatgtaattc tccttggaat
     1321 ttgccctttt tgagtttgga tcttggttca ttctcaagcc tcagacagtg gttcaaagtt
     1381 tttttcttcc atttcaggtg tcgtgactat agggggacgt cgcggccgcg tttgtttgcc
     1441 gccatggcgg aaggatccgt cgccaggcag cctgacctct tgacctgcga acatgaagag
     1501 atccacctcg ccggctcgat ccagccgcat ggcgcgcttc tggtcgtcag cgaacatgat
     1561 catcgcgtca tccaggccag cgccaacgcc gcggaatttc tgaatctcgg aagcgtactc
     1621 ggcgttccgc tcgccgagat cgacggcgat ctgttgatca agatcctgcc gcatctcgat
     1681 cccaccgccg aaggcatgcc ggtcgcggtg cgctgccgga tcggcaatcc ctctacggag
     1741 tactgcggtc tgatgcatcg gcctccggaa ggcgggctga tcatcgaact cgaacgtgcc
     1801 ggcccgtcga tcgatctgtc aggcacgctg gcgccggcgc tggagcggat ccgcacggcg
     1861 ggttcactgc gcgcgctgtg cgatgacacc gtgctgctgt ttcagcagtg caccggctac
     1921 gaccgggtga tggtgtatcg tttcgatgag caaggccacg gcctggtatt ctccgagtgc
     1981 catgtgcctg ggctcgaatc ctatttcggc aaccgctatc cgtcgtcgac tgtcccgcag
     2041 atggcgcggc agctgtacgt gcggcagcgc gtccgcgtgc tggtcgacgt cacctatcag
     2101 ccggtgccgc tggagccgcg gctgtcgccg ctgaccgggc gcgatctcga catgtcgggc
     2161 tgcttcctgc gctcgatgtc gccgtgccat ctgcagttcc tgaaggacat gggcgtgcgc
     2221 gccaccctgg cggtgtcgct ggtggtcggc ggcaagctgt ggggcctggt tgtctgtcac
     2281 cattatctgc cgcgcttcat ccgtttcgag ctgcgggcga tctgcaaacg gctcgccgaa
     2341 aggatcgcga cgcggatcac cgcgcttgag agctaagcta aattggttta gatctttggg
     2401 atacatgtta gatactaacg atcttcaagc tttgaagatg tcattgcatg aagaaaatta
     2461 tgggaaacac tgttcccgat aattacttac acccttagtg taacatatgg agatcactgt
     2521 ctccgataca gacactgtgg taggtaaaaa ctaccttgtt ctttatacat tatggaagac
     2581 actgctcccg ataatgtgtt aggattgtga caaaggtact ggaaattttg caaaatggga
     2641 attagagtta caccttggat aaactgagct ataaaatttt gcaggagtac taccacagcc
     2701 tttaagtgac attgatttat aacttggtca caattcactg catttaggaa aaccagcatt
     2761 cttatctggt cagtgctcgc ttcttagcaa cccctaatta aatttaattc atctctaaat
     2821 cttagcttca actttattca attacatttg gctgacggct gttttctaaa acccttaagt
     2881 gttgaccata aatgcaaaac ttccagtatc tgttgggttt tattagcaga tgctgctttt
     2941 atttaaaaaa aaccgacagt ataactgtca taattatgga aggcactgct tccgataatt
     3001 atattctatt aaaaaaacac catttatagt gaactctgtc actgataaat aaacaataaa
     3061 tatctcagtg ccaaaaggac agaaagctct cccctaagat taacactttg gtgtacaatc
     3121 gatgtcgaca aactacaatc ccgggcgaat agatctgtct ctgtgccttc tagttgccag
     3181 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
     3241 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
     3301 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat
     3361 gctggggatg cggtgggctc tatggtctag ccaacaagct cgtcatcgct ttgcagaaga
     3421 gcagagagga tatgctcatc gtctaaagaa ctacccattt tattatatat tagtcacgat
     3481 gtaagcatct ataacaagaa aatatatata taataagtta tcacgtaagt agaacatgaa
     3541 ataacaatat aattatcgta tgagttaaat cttaaaagtc acgtaaaaga taatcatgcg
     3601 tcattttgac tcacgcggtc gttatagttc aaaatcagtg acacttaccg cattgacaag
     3661 cacgcctcac gggagctcca agcggcgact gagatgtcct aaatgcacag cgacggattc
     3721 gcgctattta gaaagagaga gcaatatttc aagaatgcat gcgtcaattt tacgcagact
     3781 atctttctag ggttaaaaaa gatttgcgct ttactcgacc taaactttaa acacgtcata
     3841 gaatcttcgt ttgacaaaaa ccacattgtg gccaagctgt gtgacgcgac gcgcgctaaa
     3901 gaatggcaaa ccaagtcgcg cgaggtacca gcttttgttc cctttagtga gggttaattt
     3961 cgagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     4021 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     4081 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     4141 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     4201 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     4261 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     4321 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     4381 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     4441 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     4501 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     4561 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     4621 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     4681 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     4741 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     4801 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     4861 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     4921 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     4981 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     5041 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     5101 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     5161 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     5221 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     5281 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     5341 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     5401 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     5461 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     5521 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     5581 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     5641 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     5701 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     5761 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     5821 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     5881 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     5941 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     6001 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     6061 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     6121 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     6181 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     6241 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     6301 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     6361 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     6421 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     6481 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     6541 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt
     6601 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
     6661 ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc
     6721 acgacgttgt aaaacgacgg ccagtgaatt gtaatacgac tcactatagg gcgaattgga
     6781 gctcggtatt cacgacagca ggctgaataa taaaaaaatt agaaactatt atttaaccct
     6841 agaaagataa tcatattgtg acgtacgtta aagataatca tgcgtaaaat tgacgcatgt
     6901 gttttatcgg tctgtatatc gaggtttatt tattaatttg aatagatatt aagttttatt
     6961 atatttacac ttacatacta ataataaatt caacaaacaa tttatttatg tttatttatt
     7021 tattaaaaaa aaacaaaaac t
//
