LOCUS       sequence-434259-        4583 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sTb(N) half of CaST for HEK cell expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4583)
  AUTHORS   Zhang R, Anguiano M, Aarrestad IK, Lin S, Chandra J, Vadde SS, Olson
            DE, Kim CK
  TITLE     Rapid, biochemical tagging of cellular activity history in vivo.
  JOURNAL   Nat Methods. 2024 Sep;21(9):1725-1735. doi: 
            10.1038/s41592-024-02375-7. Epub 2024 Aug 5.
  PUBMED    39103446
REFERENCE   2  (bases 1 to 4583)
  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: 
            "10.1038/s41592-024-02375-7"; journalName: "Nat Methods"; date: 
            "2024-09"; volume: "21"; issue: "9"; pages: "1725-1735"
COMMENT     Sequence Label: pAAV-CMV-CaM-V5-sTb(N)
FEATURES             Location/Qualifiers
     source          1..4583
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(63..84)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     275..294
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   456..596
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   467..596
                     /label=AAV2 ITR
     polyA_signal    625..832
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     821..838
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(1097..1138)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     regulatory      1648..1657
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1670..1689)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1671..1689)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(1734..1937)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(1763..1783)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        1938..2241
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     repeat_region   2312..2441
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(2503..3091)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2583..2602)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3262..4053,4054..4122))
                     /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     3885..3904
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4123..4227)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4295..4313
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4351..4373)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4473..4492
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(join(4509..4583,1..381))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 ccgaaatcgg caaaatccct tataaatcaa aagaatagcc cgagatagag ttgagtgttg
       61 ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa
      121 aaaccgtcta tcagggcgat ggcccactac gtgaaccatc acccaaatca agttttttgg
      181 ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt
      241 gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg
      301 ctaaggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta
      361 atgcgccgct acagggcgcg tactatggtt gctttgacgt atgcggtgtg aaataccgca
      421 cagatgcgta aggagaaaat accgcatcag gcgcccctgc aggcagctgc gcgctcgctc
      481 gctcactgag gccgcccggg caaagcccgg gcgtcgggcg acctttggtc gcccggcctc
      541 agtgagcgag cgagcgcgca gagagggagt ggccaactcc atcactaggg gttcctgcgg
      601 ccgctcggtc cgcacgtggt tacctcccca gcatgcctgc tattgtcttc ccaatcctcc
      661 cccttgctgt cctgccccac cccacccccc agaatagaat gacacctact cagacaatgc
      721 gatgcaattt cctcatttta ttaggaaagg acagtgggag tggcaccttc cagggtcaag
      781 gaaggcacgg gggaggggca aacaacagat ggctggcaac tagaaggcac agtcgaggct
      841 gatcagcgag ctctagctcg aggaattctt attacagaat ctgtttagcg ttcagcagcg
      901 ggataggctc gggcagagag tagccctttc cgggcacggt gaacacgtcc acgccccagt
      961 cgcgcagagt ctgaatgtgc ttgttgatag cggccctgga catgcccagg gtttctccca
     1021 gctgttcgcc actatggaac tcgccattag ccaggagagc gatcagcttc agaggcacag
     1081 tattgtcttt gctagcggtg ctgtccaggc ccagcagggg gttggggatg ggcttgccac
     1141 ctccagagct gccaccagaa cctccagagc caccgctgcc cgagcccgag gtcgagcccg
     1201 agcccttctt cgctgtcatc atttgtacaa actcttcgta gtttacctga ccatccccat
     1261 cgatgtctgc ttccctgatc atttcatcaa cctcttcatc tgttaacttc tctccaaggt
     1321 ttgtcatcac gtggcgaagc tctgctgcac tgatgtagcc attgccatcc ttatcaaaca
     1381 caccgaacgc ttctctaatt tcttcttccg tgtccctgta tttcattttt cttgccatca
     1441 ttgtcaggaa ctcagggaag tcgattgtgc cgtcaccgtc ggcatctact tcattgatca
     1501 tgtcctgcag ctctgcttct gtggggttct gccccagaga ccgcatcacc gtccccagct
     1561 ccttggttgt tattgtccca tccccgtcct tgtcaaatag ggagaaagcc tctttaaatt
     1621 ctgcgatctg ctcttcagtc agttggtcca tggtggcaag cttgggtctc cctatagtga
     1681 gtcgtattaa tttcgataag ccagtaagca gtgggttctc tagttagcca gagagctctg
     1741 cttatataga cctcccaccg tacacgccta ccgcccattt gcgtcaatgg ggcggagttg
     1801 ttacgacatt ttggaaagtc ccgttgattt tggtgccaaa acaaactccc attgacgtca
     1861 atggggtgga gacttggaaa tccccgtgag tcaaaccgct atccacgccc attgatgtac
     1921 tgccaaaacc gcatcaccat ggtaatagcg atgactaata cgtagatgta ctgccaagta
     1981 ggaaagtccc ataaggtcat gtactgggca taatgccagg cgggccattt accgtcattg
     2041 acgtcaatag ggggcgtact tggcatatga tacacttgat gtactgccaa gtgggcagtt
     2101 taccgtaaat actccaccca ttgacgtcaa tggaaagtcc ctattggcgt tactatggga
     2161 acatacgtca ttattgacgt caatgggcgg gggtcgttgg gcggtcagcc aggcgggcca
     2221 tttaccgtaa gttatgtaac gcggaactcc atatatgggc tatgaactaa tgaccccgta
     2281 attgattact atgtacaacg cgtgcggccg caggaacccc tagtgatgga gttggccact
     2341 ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg
     2401 ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gacatgtgag caaaaggcca
     2461 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     2521 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     2581 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     2641 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     2701 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     2761 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     2821 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     2881 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     2941 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     3001 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     3061 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     3121 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     3181 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     3241 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     3301 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     3361 ggagggctta ccatctggcc ccagtgctgc aatgataccg cggcttccac gctcaccggc
     3421 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     3481 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     3541 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     3601 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     3661 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     3721 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     3781 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     3841 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     3901 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     3961 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     4021 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     4081 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     4141 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     4201 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta
     4261 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
     4321 tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt
     4381 cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg
     4441 tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg tactgagagt
     4501 gcaccataaa attgtaaacg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat
     4561 cagctcattt tttaaccaat aga
//
