LOCUS       sequence-12777-e        5427 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5427)
  AUTHORS   Shigetomi E, Kracun S, Khakh BS.
  TITLE     Monitoring astrocyte calcium microdomains with improved membrane 
            targeted GCaMP reporters.
  JOURNAL   Neuron Glia Biol. 2010 Dec 16:1-9. [Epub ahead of print]
  PUBMED    21205365
REFERENCE   2  (bases 1 to 5427)
  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: "Neuron Glia
            Biol. 2010 Dec 16:1-9. [Epub ahead of print]"
COMMENT     Sequence Label: pN1-Lck-GCaMP3
FEATURES             Location/Qualifiers
     source          1..5427
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      615..624
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(738..749,750..767,768..770,771..803,804..806,807..830,
                     831..857,858..914,915..920,921..1190,1191..1208,1209..1643,
                     1644..1646,1647..1649,1650..2093)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="MRGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWN
                     KTGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDG
                     PVLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGE
                     ELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTL
                     TYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRI
                     ELKGIDFKEDGNILGHKLEYNTRDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMR
                     SLGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDG
                     NGYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     CDS             join(738..749,750..767,768..770,771..803,804..806,807..830,
                     831..857,858..914,915..920,921..1190,1191..1208,1209..1643,
                     1644..1646,1647..1649,1650..2093)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6s
                     /translation="MRGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWN
                     KTGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDG
                     PVLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGE
                     ELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTL
                     TYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRI
                     ELKGIDFKEDGNILGHKLEYNTRDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMR
                     SLGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDG
                     NGYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      837..846
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1127..1148
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1254..1275)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1515..1534)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    2215..2336
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2252..2271)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2306..2325
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2343..2798)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2480..2501)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2692..2711
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2825..2929
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2931..3288
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2931..3122
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2954..2974)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3139..3274
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3323..4117
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3323..4117
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3377..3396)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3987..4006
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4305..4324)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4349..4396
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4725..5313
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5214..5233
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctgccacc atgggctgtg gctgcagctc aaaccctgaa gatgactgga
      661 tggagaacat tgacgtgtgt gagaactgcc attatccccc gccgaagctt cgaattgatc
      721 caccggatct cgccaccatg cggggttctc atcatcatca tcatcatggt atggctagca
      781 tgactggtgg acagcaaatg ggtcgggatc tgtacgacga tgacgataag gatctcgcca
      841 ccatggtcga ctcatcacgt cgtaagtgga ataagacagg tcacgcagtc agagctatag
      901 gtcggctgag ctcactcgag aacgtctata tcaaggccga caagcagaag aacggcatca
      961 aggcgaactt caagatccgc cacaacatcg aggacggcgg cgtgcagctc gcctaccact
     1021 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
     1081 gcgtccagtc caaactttcg aaagacccca acgagaagcg cgatcacatg gtcctgctgg
     1141 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag ggcggtaccg
     1201 gagggagcat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     1261 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggt gagggcgatg
     1321 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     1381 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     1441 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacatc caggagcgca
     1501 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     1561 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     1621 tggggcacaa gctggagtac aacacgcgtg accaactgac tgaagagcag atcgcagaat
     1681 ttaaagaggc tttctcccta tttgacaagg acggggatgg gacaataaca accaaggagc
     1741 tggggacggt gatgcggtct ctggggcaga accccacaga agcagagctg caggacatga
     1801 tcaatgaagt agatgccgac ggtgatggca caatcgactt ccctgagttc ctgacaatga
     1861 tggcaagaaa aatgaaagac acagacagtg aagaagaaat tagagaagcg ttccgtgtgt
     1921 ttgataagga tggcaatggc tacatcagtg cagcagagct tcgccacgtg atgacaaacc
     1981 ttggagagaa gttaacagat gaagaggttg atgaaatgat cagggaagca gacatcgatg
     2041 gggatggtca ggtaaactac gaagagtttg tacaaatgat gacagcgaag tgagcggccg
     2101 cgactctaga tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac
     2161 ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg
     2221 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     2281 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaaggc
     2341 gtaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     2401 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     2461 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     2521 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     2581 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     2641 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     2701 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     2761 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa
     2821 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     2881 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga
     2941 aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     3001 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     3061 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     3121 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     3181 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     3241 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga
     3301 gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc
     3361 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     3421 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     3481 gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac
     3541 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     3601 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     3661 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     3721 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     3781 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     3841 gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     3901 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     3961 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     4021 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     4081 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg
     4141 accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat
     4201 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     4261 gatctcatgc tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag
     4321 acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt
     4381 gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc
     4441 ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc
     4501 ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata
     4561 gcctcaggtt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     4621 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     4681 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     4741 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     4801 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     4861 ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     4921 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     4981 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     5041 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     5101 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     5161 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     5221 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     5281 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     5341 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     5401 gtggataacc gtattaccgc catgcat
//
