LOCUS       sequence-382343-        7360 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression vector for a low affinity calcium indicator 
            exposed to the cell surface.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7360)
  AUTHORS   Valiente-Gabioud AA, Garteizgogeascoa Suner I, Idziak A, Fabritius 
            A, Basquin J, Angibaud J, Nagerl UV, Singh SP, Griesbeck O
  TITLE     Fluorescent sensors for imaging of interstitial calcium.
  JOURNAL   Nat Commun. 2023 Oct 5;14(1):6220. doi: 10.1038/s41467-023-41928-w.
  PUBMED    37798285
REFERENCE   2  (bases 1 to 7360)
  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: "2023-10-5"; volume: "14(1):6220. doi"; pages: " 
            10.1038/s41467-023-41928-w"
COMMENT     Sequence Label: GreenT-EC - Display - CyRFP1
FEATURES             Location/Qualifiers
     source          1..7360
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(118..137)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        309..688
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        689..892
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     843..863
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     937..956
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        937..955
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     sig_peptide     981..1040
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             1053..1076
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      1912..1921
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2088..2234
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(3025..3042)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3031..3255
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3301..3729
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3388..3407)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3598..3619
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3738..3758)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3743..4072
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3923..4058
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3985..4004
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4139..4933
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4193..4212)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4803..4822
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5107..5228
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5144..5163)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5198..5217
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5277..5293)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5277..5293)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5290..5312)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5301..5317
                     /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(5325..5331,5332..5349,5350..5355))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5370..5391
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5508..5525)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5679..6267)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5759..5778)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6438..7229,7230..7298))
                     /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     7061..7080
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7299..7360,1..43))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
       61 tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac
      121 aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt
      181 cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg
      241 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata
      301 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
      361 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      421 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      481 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      541 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      601 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      661 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
      721 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
      781 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
      841 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct
      901 aactagagaa cccactgctt actggcttat cgaaattaat acgactcact atagggagac
      961 ccaagctggc tagcaccgcc atggagacag acacactcct gctatgggta ctgctgctct
     1021 gggttccagg ttccactggt gacggatccg gtgattacaa ggacgacgat gacaagggtg
     1081 gtaccggtgg ctctggtagc agtggcggta gcatggtctc caagggggag gaggacaaca
     1141 tggggagcct gccagctacc cacgagctgc atatctttgg atctattaac ggggtggatt
     1201 tcgacatggt tggacaaggt agcgggaatc ctaatgatgg gtatgaagag cttaacctta
     1261 aaagcactaa gggcgatctg caattttcac cttggattct cgtcccccac attgggtatg
     1321 ggttccatca atatctccct tacccggatg gaatgtctcc cttccaagcg gctatggtag
     1381 atggtagtgg ttaccaagtt cacaggacta tgcaattcga agacggcgcc tcattgactg
     1441 tgaactaccg atacacttac gaaggatctc acataaaagg tgaagcgcag gtgaagggca
     1501 cgggatttcc agcagatggg ccggtaatga ctaactcact tactgcggca gacttggggt
     1561 gggattcaga agaggaactt tccgaatgtt ttagaatttt tgattttgac ggtaacgacg
     1621 gattcattga ccgcgaagag ttcggtgaca taatcagatt gactggggag caacttacgg
     1681 atgaggacgt agacgaaatt tttggcgact cagatacgga taagaatggc cggatcgact
     1741 ttgatgaatt tctgaaaatg gtcgaaaatg ttcagctcac ggacaaccga acaacgaaaa
     1801 agacctatcc aaatgacaag acaattatct caactttgaa atggagctac accacggaca
     1861 acggcaagag gtatcggtcc accgctatca ctacttatac atttgctaaa cccatggcgg
     1921 ctaactacct caagaatcaa cctatgtacg tgtttcgcaa gactgagctt aaacatagca
     1981 aaaccgaatt gaatttcaag gagtggcaga aggcgtttac ggacgtaatg gggatggatg
     2041 agctttataa gggtggtacc ggtggctctg gtagcagtgg cggtagcgct gtgggccagg
     2101 acacgcagga ggtcatcgtg gtgccacact ccttgccctt taaggtggtg gtgatctcag
     2161 ccatcctggc cctggtggtg ctcaccatca tctcccttat catcctcatc atgctttggc
     2221 agaagaagcc acgtggtggt actggcggta gcggttctag tggcggttct gatatcatgg
     2281 tgagcaaggg cgaggagctg atcaaggaga acatgagaag caagctgtac ctggaaggca
     2341 gcgtgaacgg ccaccagttc aagtgcaccc acgaagggga gggcaagccc tacgagggca
     2401 agcagaccgc caggatcaag gtggtggagg gaggccccct gccgttcgca ttcgacatcc
     2461 tggccaccat gtttatgtac gggagcaagg tgttcatcaa gtaccccgcc gacctccccg
     2521 attattttaa gcagtccttc cctgagggct tcacatggga gagagtcatg gtgttcgaag
     2581 acgggggcgt gctgaccgcc acccaggaca ccagcctcca ggacggcgag ctcatctaca
     2641 acgtcaagct cagaggggtg aacttcccag ccaacggccc cgtgatgcag aagaaaacac
     2701 tgggctggga gcccagcacc gagaccatgt accccgctga cggcggcctg gaaggcagat
     2761 gcgacaagaa gctgaagctc gtgggcgggg gccacctgca cgtcaacttc aagaccacat
     2821 acaagtccaa gaaacccgtg aagatgcccg gcgtccacta cgtggaccgc agactggaaa
     2881 gaatcaagga ggccgacaac gagacctacg tcgagcagta cgagcacgct gtggccagat
     2941 actccaacct gggcggaggc atggacgagc tgtaaagtaa gcggccgctc gagtctagag
     3001 ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
     3061 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     3121 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     3181 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
     3241 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
     3301 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     3361 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     3421 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     3481 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     3541 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     3601 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     3661 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     3721 acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     3781 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     3841 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     3901 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     3961 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga
     4021 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc
     4081 gggagcttgt atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat
     4141 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg
     4201 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc
     4261 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca
     4321 ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct
     4381 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga
     4441 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg
     4501 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat
     4561 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga
     4621 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg
     4681 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg
     4741 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat
     4801 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct
     4861 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga
     4921 cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg
     4981 ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     5041 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     5101 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     5161 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     5221 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
     5281 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     5341 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     5401 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     5461 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     5521 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     5581 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     5641 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     5701 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     5761 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     5821 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     5881 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     5941 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     6001 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     6061 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     6121 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     6181 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     6241 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     6301 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     6361 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     6421 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     6481 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     6541 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     6601 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     6661 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     6721 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     6781 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     6841 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     6901 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     6961 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     7021 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     7081 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     7141 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     7201 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     7261 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     7321 tgaagcattt atcagggtta ttgtctcatg agcggataca
//
