LOCUS       sequence-199364-        7904 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Presynaptic calcium sensor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7904)
  AUTHORS   Kwon SK, Sando R 3rd, Lewis TL, Hirabayashi Y, Maximov A, Polleux F
  TITLE     LKB1 Regulates Mitochondria-Dependent Presynaptic Calcium Clearance 
            and Neurotransmitter Release Properties at Excitatory Synapses along
            Cortical Axons.
  JOURNAL   PLoS Biol. 2016 Jul 18;14(7):e1002516. doi: 
            10.1371/journal.pbio.1002516. eCollection 2016 Jul.
  PUBMED    27429220
REFERENCE   2  (bases 1 to 7904)
  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: "PLoS 
            Biol."; date: "2016-07-18"; volume: "14(7):e1002516. doi"; pages: " 
            10.1371/journal.pbio.1002516. eCollection 2016 Jul"
COMMENT     Sequence Label: pCAG vGlut1 GCaMP5G
FEATURES             Location/Qualifiers
     source          1..7904
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(42..629)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(122..141)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(800..1591,1592..1660))
                     /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     1423..1442
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1661..1765)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        1796..2175
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2177..2454
                     /label=chicken beta-actin promoter
     intron          2455..3467
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     3391..3413
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     3475..3494
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      3544..3553
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(4835..4846)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             join(5245..5253,5254..5271,5272..5274,5275..5307,5308..5310,
                     5311..5334,5335..5361,5362..5418,5419..5424,5425..5694,
                     5695..5712,5713..6147,6148..6153,6154..6597)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYTDSEEEIREAFRVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     CDS             join(5245..5253,5254..5271,5272..5274,5275..5307,5308..5310,
                     5311..5334,5335..5361,5362..5418,5419..5424,5425..5694,
                     5695..5712,5713..6147,6148..6153,6154..6597)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6s
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYTDSEEEIREAFRVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      5341..5350
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     5631..5652
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(5758..5779)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(6019..6038)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     complement(6688..6707)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    6753..6808
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(6807..6826)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(7199..7215)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7199..7215)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7212..7234)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7223..7239
                     /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(7247..7253,7254..7271,7272..7277))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7292..7313
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        7372..7567
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      7418..7553
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    7573..7707
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7610..7629)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7664..7683
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(7775..7792)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
       61 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
      121 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
      181 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
      241 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
      301 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
      361 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
      421 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
      481 agacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
      541 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
      601 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
      661 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
      721 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
      781 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
      841 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
      901 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
      961 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     1021 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     1081 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     1141 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     1201 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     1261 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     1321 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     1381 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     1441 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     1501 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     1561 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     1621 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     1681 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     1741 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gggtcgacat
     1801 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
     1861 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
     1921 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
     1981 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     2041 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     2101 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
     2161 atcgctatta ccatggtcga ggtgagcccc acgttctgct tcactctccc catctccccc
     2221 ccctccccac ccccaatttt gtatttattt attttttaat tattttgtgc agcgatgggg
     2281 gcgggggggg ggggggggcg cgcgccaggc ggggcggggc ggggcgaggg gcggggcggg
     2341 gcgaggcgga gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa gtttcctttt
     2401 atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg ggcgggagtc
     2461 gctgcgttgc cttcgccccg tgccccgctc cgcgccgcct cgcgccgccc gccccggctc
     2521 tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc tccgggctgt
     2581 aattagcgct tggtttaatg acggctcgtt tcttttctgt ggctgcgtga aagccttaaa
     2641 gggctccggg agggcccttt gtgcgggggg gagcggctcg gggggtgcgt gcgtgtgtgt
     2701 gtgcgtgggg agcgccgcgt gcggcccgcg ctgcccggcg gctgtgagcg ctgcgggcgc
     2761 ggcgcggggc tttgtgcgct ccgcgtgtgc gcgaggggag cgcggccggg ggcggtgccc
     2821 cgcggtgcgg gggggctgcg aggggaacaa aggctgcgtg cggggtgtgt gcgtgggggg
     2881 gtgagcaggg ggtgtgggcg cggcggtcgg gctgtaaccc ccccctgcac ccccctcccc
     2941 gagttgctga gcacggcccg gcttcgggtg cggggctccg tacggggcgt ggcgcggggc
     3001 tcgccgtgcc gggcgggggg tggcggcagg tgggggtgcc gggcggggcg gggccgcctc
     3061 gggccgggga gggctcgggg gaggggcgcg gcggcccccg gagcgccggc ggctgtcgag
     3121 gcgcggcgag ccgcagccat tgccttttat ggtaatcgtg cgagagggcg cagggacttc
     3181 ctttgtccca aatctgtgcg gagccgaaat ctgggaggcg ccgccgcacc ccctctagcg
     3241 ggcgcggggc gaagcggtgc ggcgccggca ggaaggaaat gggcggggag ggccttcgtg
     3301 cgtcgccgcg ccgccgtccc cttctccctc tccagcctcg gggctgtccg cggggggacg
     3361 gctgccttcg ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg accggcggct
     3421 ctagagcctc tgctaaccat gttcatgcct tcttcttttt cctacagctc ctgggcaacg
     3481 tgctggttat tgtgctgtct catcattttg gcaaagaatt gctcgagctc aagcttcgaa
     3541 ttggccacca tggagttccg gcaggaggag tttcggaagc tggcggggcg cgccctgggg
     3601 aggctgcacc ggttactgga gaagcggcag gaaggcgcgg agacattgga gctgagcgcc
     3661 gacgggcgcc cagtgaccac acacacgcgg gacccgccgg tggtggactg cacttgcttt
     3721 ggcctccctc gccgctacat catcgcgatc atgagcggtc tgggtttctg catcagcttt
     3781 ggcatccgct gcaacctggg cgtggccatc gtatccatgg tcaacaacag tacaacccac
     3841 cgtgggggcc acgtggtggt gcagaaagcc cagttcaact gggatccaga gactgtcggc
     3901 ctcatacatg gctccttttt ctgggggtac attgtcactc agattcctgg aggatttatc
     3961 tgccaaaaat tcgcagccaa cagggtcttt ggctttgcca ttgtggctac ctccacccta
     4021 aatatgttga tcccttcagc agcccgtgtt cactatggct gtgtcatctt cgtgaggatc
     4081 cttcagggat tggtggaggg ggtcacatac cctgcttgcc atggcatctg gagcaaatgg
     4141 gcccctccct tagaacggag tcggctggcg acgacagcct tttgcggttc ctatgccggg
     4201 gcagtggttg ccatgcctct ggctggggtc ctggtacagt attcaggatg gagttctgtc
     4261 ttctatgtct atggcagctt cgggatcttt tggtacctgt tctggttgct tgtctcctac
     4321 gagtcacctg cactacaccc cagcatctcc gaggaggagc gcaaatacat tgaggatgcc
     4381 atcggagaaa gcgccaagct catgaaccct gttacgaagt ttaacacacc ctggaggcgc
     4441 ttctttacct ccatgccggt ctatgccatc attgtcgcca acttttgccg cagctggact
     4501 ttctacctgc tcctcatctc ccagcccgcc tactttgaag aagtgttcgg ctttgagatc
     4561 agcaaggtgg gactggtgtc ggcactgcct caccttgtca tgactatcat cgtacccatc
     4621 ggaggccaga tcgccgactt cctgcgcagt cgtcatataa tgtccacgac caatgtgcga
     4681 aagctgatga actgcggggg tttcgggatg gaagctacgc tgctgctggt ggtcggatac
     4741 tcacactcca agggcgtggc catctccttc ctggtcctgg ctgtgggctt cagtggcttt
     4801 gctatctctg ggtttaacgt gaaccacttg gacatcgccc ctcgatatgc cagcatcttg
     4861 atgggcattt ccaatggcgt gggcacactg tctgggatgg tgtgccccat catcgtgggt
     4921 gcaatgacca agcacaagac gcgggaggag tggcagtacg tgttcctcat agcctccctg
     4981 gtgcactatg gaggtgtcat cttctatggg gtctttgctt cgggagagaa acagccgtgg
     5041 gcagagccgg aggagatgag cgaggagaag tgtggctttg ttggccacga ccagctggct
     5101 ggcagtgacg aaagtgaaat ggaagacgag gttgagcccc cgggggcacc ccccgcacct
     5161 ccgccttcct acggggccac acacagcaca gttcagcctc caaggccccc accccctgtc
     5221 cgggactacg aattcggtgg ttctatgggt tctcatcatc atcatcatca tggtatggct
     5281 agcatgactg gtggacagca aatgggtcgg gatctgtacg acgatgacga taaggatctc
     5341 gccaccatgg tcgactcatc acgtcgtaag tggaataaga caggtcacgc agtcagagct
     5401 ataggtcggc tgagctcact cgagaacgtc tatatcaagg ccgacaagca gaagaacggc
     5461 atcaaggcga acttcaagat ccgccacaac atcgaggacg gcggcgtgca gctcgcctac
     5521 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     5581 ctgagcgtgc agtccaaact ttcgaaagac cccaacgaga agcgcgatca catggtcctg
     5641 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagggcggt
     5701 accggaggga gcatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     5761 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggtgagggc
     5821 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     5881 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
     5941 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta catccaggag
     6001 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     6061 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     6121 atcctggggc acaagctgga gtacaacctg ccggaccaac tgactgaaga gcagatcgca
     6181 gaatttaaag aggctttctc cctatttgac aaggacgggg atgggacaat aacaaccaag
     6241 gagctgggga cggtgatgcg gtctctgggg cagaacccca cagaagcaga gctgcaggac
     6301 atgatcaatg aagtagatgc cgacggtgac ggcacaatcg acttccctga gttcctgaca
     6361 atgatggcaa gaaaaatgaa atacacagac agtgaagaag aaattagaga agcgttccgt
     6421 gtgtttgata aggatggcaa tggctacatc agtgcagcag agcttcgcca cgtgatgaca
     6481 aaccttggag agaagttaac agatgaagag gttgatgaaa tgatcaggga agcagacatc
     6541 gatggggatg gtcaggtaaa ctacgaagag tttgtacaaa tgatgacagc gaagtgagcg
     6601 gccgcaattc actcctcagg tgcaggctgc ctatcagaag gtggtggctg gtgtggccaa
     6661 tgccctggct cacaaatacc actgagatct ttttccctct gccaaaaatt atggggacat
     6721 catgaagccc cttgagcatc tgacttctgg ctaataaagg aaatttattt tcattgcaat
     6781 agtgtgttgg aattttttgt gtctatcact cggaaggaca tatgggaggg caaatcattt
     6841 aaaacatcag aatgagtatt tggtttagag tttggcaaca tatgcccata tgctggctgc
     6901 catgaacaaa ggttggctat aaagaggtca tcagtatatg aaacagcccc ctgctgtcca
     6961 ttccttattc catagaaaag ccttgacttg aggttagatt ttttttatat tttgttttgt
     7021 gttatttttt tctttaacat ccctaaaatt ttccttacat gttttactag ccagattttt
     7081 cctcctctcc tgactactcc cagtcatagc tgtccctctt ctcttatgga gatccctcga
     7141 cctgcaccgt cgaccagctg gtcgacggtg caccgtcgac cagcttggcg taatcatggt
     7201 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     7261 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     7321 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagcggatc cgcatctcaa
     7381 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     7441 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     7501 cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     7561 ttgcaaaaag ctaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     7621 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     7681 atcaatgtat cttatcatgt ctggatccgc tgcattaatg aatcggccaa cgcgcgggga
     7741 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     7801 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     7861 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccag
//
