LOCUS       sequence-492101-        7105 bp    DNA     circular SYN 06-NOV-2025
DEFINITION  Cre dependent bioluminescent reporter for calcium signaling.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7105)
  AUTHORS   Lambert GG, Crespo EL, Murphy J, Boassa D, Luong S, Hu J, Sprecher 
            B, Tree MO, Orcutt R, Heydari D, Bell AB, Torreblanca Zanca A, 
            Hakimi A, Lipscombe D, Moore CI, Hochgeschwender U, Shaner NC
  TITLE     CaBLAM! A high-contrast bioluminescent Ca (2+) indicator derived 
            from an engineered Oplophorus gracilirostris luciferase.
  JOURNAL   bioRxiv. 2023 Jun 26:2023.06.25.546478. doi: 
            10.1101/2023.06.25.546478. Preprint.
  PUBMED    37425712
REFERENCE   2  (bases 1 to 7105)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 6, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv. 
            2023 Jun 26:2023.06.25.546478. doi: 10.1101/2023.06.25.546478. 
            Preprint."
COMMENT     Sequence Label: pAAV EfIa DIO CaBLAM
FEATURES             Location/Qualifiers
     source          1..7105
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        242..1420
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          473..1411
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1368..1388
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1451..1484)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(1535..1568)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(2139..2195)
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKTGHAVRAIGRLSS"
     regulatory      3095..3104
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      3356..3365
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3382..3415
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    3466..3499
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3524..4112
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3577..3597)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3995..4006)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4145..4256
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4368..4508
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4368..4497
                     /label=AAV2 ITR
     rep_origin      4583..5038
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4670..4689)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4880..4901
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5055..5074)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5174..5196
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5234..5252)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5320..5424
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5425..5493,5494..6285)
                     /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     complement(5643..5662)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6456..7044
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6945..6964
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt
      181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
      241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
      301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
      361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
      421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc
      481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat
      541 tacttccacc tggctgcagt acgtgattct tgatcccgag cttcgggttg gaagtgggtg
      601 ggagagttcg aggccttgcg cttaaggagc cccttcgcct cgtgcttgag ttgaggcctg
      661 gcctgggcgc tggggccgcc gcgtgcgaat ctggtggcac cttcgcgcct gtctcgctgc
      721 tttcgataag tctctagcca tttaaaattt ttgatgacct gctgcgacgc tttttttctg
      781 gcaagatagt cttgtaaatg cgggccaaga tctgcacact ggtatttcgg tttttggggc
      841 cgcgggcggc gacggggccc gtgcgtccca gcgcacatgt tcggcgaggc ggggcctgcg
      901 agcgcggcca ccgagaatcg gacgggggta gtctcaagct ggccggcctg ctctggtgcc
      961 tggcctcgcg ccgccgtgta tcgccccgcc ctgggcggca aggctggccc ggtcggcacc
     1021 agttgcgtga gcggaaagat ggccgcttcc cggccctgct gcagggagct caaaatggag
     1081 gacgcggcgc tcgggagagc gggcgggtga gtcacccaca caaaggaaaa gggcctttcc
     1141 gtcctcagcc gtcgcttcat gtgactccac ggagtaccgg gcgccgtcca ggcacctcga
     1201 ttagttctcg agcttttgga gtacgtcgtc tttaggttgg ggggaggggt tttatgcgat
     1261 ggagtttccc cacactgagt gggtggagac tgaagttagg ccagcttggc acttgatgta
     1321 attctccttg gaatttgccc tttttgagtt tggatcttgg ttcattctca agcctcagac
     1381 agtggttcaa agtttttttc ttccatttca ggtgtcgtga ggtaccggat cctctagagt
     1441 cgactccgga ataacttcgt ataggatact ttatacgaag ttatgcagaa tggtagctgg
     1501 attgtagctg ctattagcaa tatgaaacct cttaataact tcgtatagca tacattatac
     1561 gaagttatgg cgcgccacgc gtgcatatgt tactcgaggt tagagatttc ttcgagcaga
     1621 cggtaaccgg taacagcagg agacttagcg gtcatcatct gaacgaattc ttcgtagtta
     1681 acctgaccgt caccgtcgat gtcagcttca cggatcattt cgtcaacttc ttcgtcggtc
     1741 agcttttcac ccaggttggt cataacatga cgcagttcag cagcagagat gtaaccgata
     1801 ccgtccttgt cgaaaacgcc gaaagcttca cggatttcct cttcggtgtc cctgtacttc
     1861 atcttacgag ccatcatggt caggaattca gggaagtcga tggtaccgtc accgtcagcg
     1921 tcaacttcgt tgatcatgtc ttgcagttca gcttcggtag ggttctgacc cagagaacgc
     1981 atcacggtac ccagttcctt ggtggtgatg gtaccgtctc catctttatc gaacagagag
     2041 aactcttcct tgaattcagc gatctgttct tcggtcagct ggtcaggcag gttggatcca
     2101 ccagaacctc cggatccacc actaccaccg gatccaccag aagacagacg accgatagca
     2161 cgaacagcat gaccggtctt gttccactta cgcctagaag agtcagaaga agcagagttg
     2221 atggtaacac ggaacagcat agaaccctca ggggtgatca gacgttcgtc gatgatcttg
     2281 ttaccgttcc acagggtacc ggtaacggtg atcttcttac cgtcgaaaac agcgatacct
     2341 tcgtaaggac gaccgaagta gttcagcatg ttaggggtaa caccgtcgat aaccagggta
     2401 ccgtattcca tgataacctt gaaatgatgg tcgtcaacag ggtaaacaac cttgaaaact
     2461 tcttcgatat gagccatctg gtcagcagac agaccttcgt aagggatgat aacatggatg
     2521 tcgatcttca gaccgttttc accagaacga acgatacgct ggataggggt aacagaaaca
     2581 gccagggtct gcagaacaga agaaacacca ccctgttcca gaacctggtc caggttgtaa
     2641 gcagcggtct gttcccagtc accaacgaag tcttcaaatc cggtaaaggc cttttgccac
     2701 tccttgaagt tgagctcggt cttggagtgc ttgagctccg tcttacggaa cacgtacatc
     2761 ggctggttct tcagatagtt agccgccatt ggcttggcaa aggtgtaggt ggtccgcgca
     2821 gtgctccggt agcgcttgcc atttccagtg gtgtaactcc acttaaaggt actgatgatg
     2881 gttttgtcgt tggggtaagt cttcttcgac ctgcaccagt ccgcagcggt cagcgagttg
     2941 gtcatcacag gaccgtcagc agggaaacca gtccccttca cctgggcctc tcctttgatg
     3001 tggcttccct cgtaggtgta gcggtagtta acagtaaggg aggcaccatc ttcaaactgc
     3061 attgtgcgat ggacttggta tccggagcca tctaccatgg cggcctggaa aggcgacatc
     3121 ccgtcagggt agggcaggta ctgatggaag ccatacccga tatgagggac cagaatccag
     3181 ggggagaact ggaggtcacc cttggtggac ttcaggttta actcctcata accatcattt
     3241 ggattgccgg tgccctgacc caccatgtca aagtccacac cgttgatgga gccaaagatg
     3301 tgtaactcat gtgtcgctgg gagagaggcc atgttatcct cctcgccctt gctcaccatg
     3361 gtggcgaccg gtagcgctag cataacttcg tataaagtat cctatacgaa gttatttgcc
     3421 ttaacccaga aattatcact gttattcttt agaatggtgc aaagaataac ttcgtataat
     3481 gtatgctata cgaagttatg aattcgatat caagcttatc gataatcaac ctctggatta
     3541 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg
     3601 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc
     3661 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca
     3721 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac
     3781 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact
     3841 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     3901 cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctatg ttgccacctg
     3961 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc
     4021 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac
     4081 gagtcggatc tccctttggg ccgcctcccc gcatcgatac cgagcgctgc tcgagagatc
     4141 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     4201 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
     4261 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     4321 gattgggaag acaatagcag gcatgcacgt gcggaccgag cggccgcagg aacccctagt
     4381 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
     4441 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg
     4501 cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     4561 catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     4621 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     4681 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     4741 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     4801 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     4861 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     4921 tcgggctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     4981 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta
     5041 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     5101 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     5161 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     5221 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     5281 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     5341 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     5401 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     5461 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     5521 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     5581 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     5641 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     5701 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     5761 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     5821 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     5881 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     5941 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     6001 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     6061 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     6121 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     6181 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     6241 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     6301 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     6361 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     6421 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     6481 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     6541 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     6601 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     6661 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     6721 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     6781 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6841 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6901 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6961 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     7021 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     7081 ttttgctggc cttttgctca catgt
//
