LOCUS       sequence-362800-        4680 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses nuclear-localized LgBiT-FRB.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4680)
  AUTHORS   Erdenee E, Ting AY
  TITLE     A Dual-Purpose Real-Time Indicator and Transcriptional Integrator 
            for Calcium Detection in Living Cells.
  JOURNAL   ACS Synth Biol. 2022 Mar 18;11(3):1086-1095. doi: 
            10.1021/acssynbio.1c00597. Epub 2022 Mar 7.
  PUBMED    35254056
REFERENCE   2  (bases 1 to 4680)
  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"; doi: 
            "10.1021/acssynbio.1c00597"; journalName: "ACS Synth Biol"; date: 
            "2022-03-18- 18"; volume: "11"; issue: "3"; pages: "1086-1095"
COMMENT     Sequence Label: NLS-LgBiT-FRB
FEATURES             Location/Qualifiers
     source          1..4680
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        201..504
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        505..708
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     659..679
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     753..772
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        753..771
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             827..847
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1379..1402
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(1701..1718)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1707..1914
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   1943..2083
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   1943..2072
                     /label=AAV2 ITR
     rep_origin      2158..2613
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2245..2264)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2455..2476
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2630..2649)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2749..2771
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2809..2827)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2895..2999
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3000..3068,3069..3860)
                     /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(3218..3237)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4031..4619
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4520..4539
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgttgtaca tagtaatcaa ttacggggtc attagttcat
      181 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      241 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      301 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      361 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      421 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      481 gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga
      541 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
      601 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
      661 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact
      721 agagaaccca ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa
      781 gcttgccacc atgaagcgga ctgctgatgg cagtgaattt gagagcccca aaaagaagag
      841 gaaagtggtc ttcactctgg aagatttcgt aggtgactgg gagcaaacag cagcatacaa
      901 tcttgatcaa gtgttggagc aagggggggt aagctctctc ctccaaaact tggctgtctc
      961 agttactcca attcaacgaa ttgtacggtc aggggaaaat gcactcaaga ttgacattca
     1021 tgtcatcatt ccttatgagg gtctgagtgc cgatcagatg gcccagatcg aggaggtttt
     1081 taaggtagtg tatcccgtgg atgatcatca ctttaaggta attttgccct acggaactct
     1141 ggtaattgac ggagtgaccc ccaatatgct caattatttt ggtaggccct atgaaggtat
     1201 tgctgtcttc gacggaaaaa aaataacagt tactgggaca ctctggaatg gtaataagat
     1261 tatagacgaa cgcttgatca ctcctgacgg cagtatgttg ttccgcgtta ccataaatgg
     1321 gtctgggaac ggaaatggca atggtaacgg gaatggaaac ggcaatggta acgggaatga
     1381 ctacaaggat gacgacgata agatgtggca tgaaggcctg gaagaggcat ctcgtttgta
     1441 ctttggggaa aggaacgtga aaggcatgtt tgaggtgctg gagcccttgc atgctatgat
     1501 ggaacggggc ccccagactc tgaaggaaac atcctttaat caggcctatg gtcgagattt
     1561 aatggaggcc caagagtggt gcaggaagta catgaaatca gggaatgtca aggacctcac
     1621 ccaagcctgg gacctctatt atcatgtgtt ccgacgaatc tcaaagtagg aattcctcga
     1681 gctagagcct cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     1741 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     1801 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     1861 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggaggtaac
     1921 cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
     1981 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     2041 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat
     2101 tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg
     2161 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     2221 cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     2281 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     2341 ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat
     2401 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     2461 tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt gatttataag
     2521 ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     2581 cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct
     2641 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     2701 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     2761 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     2821 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     2881 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     2941 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     3001 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     3061 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     3121 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     3181 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     3241 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     3301 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     3361 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     3421 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     3481 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     3541 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     3601 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     3661 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtggaagcc
     3721 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     3781 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     3841 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     3901 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     3961 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     4021 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     4081 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     4141 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     4201 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     4261 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     4321 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     4381 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     4441 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     4501 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     4561 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     4621 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
//
