LOCUS       sequence-178498-        5770 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  pPBC-LG6f expresses PiggyBac inverted terminal repeat-flanked 
            Lck-GCaMP6f under the control of the CAG promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5770)
  AUTHORS   Gee JM, Gibbons MB, Taheri M, Palumbos S, Morris SC, Smeal RM, Flynn
            KF, Economo MN, Cizek CG, Capecchi MR, Tvrdik P, Wilcox KS, White JA
  TITLE     Imaging activity in astrocytes and neurons with genetically encoded 
            calcium indicators following in utero electroporation.
  JOURNAL   Front Mol Neurosci. 2015 Apr 15;8:10. doi: 10.3389/fnmol.2015.00010.
            eCollection 2015.
  PUBMED    25926768
REFERENCE   2  (bases 1 to 5770)
  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: "Front Mol 
            Neurosci."; date: "2015-04-15"; volume: "8:10. doi"; pages: " 
            10.3389/fnmol.2015.00010. eCollection 2015"
COMMENT     Sequence Label: pPBC-LG6f
FEATURES             Location/Qualifiers
     source          1..5770
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     39..61
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     123..142
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      188..197
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(311..322,323..340,341..343,344..376,377..379,380..403,
                     404..430,431..487,488..493,494..763,764..781,782..1216,
                     1217..1222,1223..1666)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="MRGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWN
                     KTGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDG
                     PVLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGE
                     ELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTL
                     TYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRI
                     ELKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEEFSLFDKDGDGTITTKELGTVMR
                     SLGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDG
                     NGYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      410..419
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     700..721
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(827..848)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1088..1107)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     complement(1848..1867)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1892..1939
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     repeat_region   2287..2321
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(2445..2464)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2446..2464)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2551..2568)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2722..3310)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2802..2821)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3481..4272,4273..4341))
                     /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     4104..4123
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4342..4446)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4514..4532
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4570..4592)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4692..4711
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4792..4810
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4792..4809
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    4876..4897
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4912..4917,4918..4935,4936..4942)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4950..4966
                     /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)."
     primer_bind     4955..4977
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4974..4990
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4974..4990
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        5011..5029
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     repeat_region   5212..5274
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
ORIGIN
        1 gggggacggc tgccttcggg ggggacgggg cagggcgggg ttcggcttct ggcgtgtgac
       61 cggcggctct agagcctctg ctaaccatgt tcatgccttc ttctttttcc tacagctcct
      121 gggcaacgtg ctggttattg tgctgtctca tcattttggc aaagctagcg ctaccggact
      181 cagatctgcc accatgggct gtggctgcag ctcaaaccct gaagatgact ggatggagaa
      241 cattgacgtg tgtgagaact gccattatcc cccgccgaag cttcgaattg atccaccgga
      301 tctcgccacc atgcggggtt ctcatcatca tcatcatcat ggtatggcta gcatgactgg
      361 tggacagcaa atgggtcggg atctgtacga cgatgacgat aaggatctcg ccaccatggt
      421 cgactcatca cgtcgtaagt ggaataagac aggtcacgca gtcagagcta taggtcggct
      481 gagctcactc gagaacgtct atatcaaggc cgacaagcag aagaacggca tcaaggcgaa
      541 cttcaagatc cgccacaaca tcgaggacgg cggcgtgcag ctcgcctacc actaccagca
      601 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcgtgca
      661 gtccaaactt tcgaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
      721 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagggcggta ccggagggag
      781 catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
      841 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggtgagggcg atgccaccta
      901 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
      961 cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa
     1021 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac atccaggagc gcaccatctt
     1081 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
     1141 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
     1201 caagctggag tacaacctgc cggaccaact gactgaagag cagatcgcag aatttaaaga
     1261 ggaattctcc ctatttgaca aggacgggga tgggacaata acaaccaagg agctggggac
     1321 ggtgatgcgg tctctggggc agaaccccac agaagcagag ctgcaggaca tgatcaatga
     1381 agtagatgcc gacggtgacg gcacaatcga cttccctgag ttcctgacaa tgatggcaag
     1441 aaaaatgaaa tacagggaca cggaagaaga aattagagaa gcgttcggtg tgtttgataa
     1501 ggatggcaat ggctacatca gtgcagcaga gcttcgccac gtgatgacaa accttggaga
     1561 gaagttaaca gatgaagagg ttgatgaaat gatcagggaa gcagacatcg atggggatgg
     1621 tcaggtaaac tacgaagagt ttgtacaaat gatgacagcg aagtgagcgg ccgcttcgaa
     1681 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc
     1741 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     1801 ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga aacacggaag
     1861 gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa taaaacgcac
     1921 ggtgttgggt cgtttgttcg gcgcgccatc gaagcagctc cagcctacac aatcgctcaa
     1981 gacgtgtaat gcttttatta tatattagtc acgatatcta taacaagaaa atatatatat
     2041 aataagttat cacgtaagta gaacatgaaa taacaatata attatcgtat gagttaaatc
     2101 ttaaaagtca cgtaaaagat aatcatgcgt cattttgact cacgcggtcg ttatagttca
     2161 aaatcagtga cacttaccgc attgacaagc acgcctcacg ggagctccaa gcggcgactg
     2221 agatgtccta aatgcacagc gacggattcg cgctatttag aaagagagag caatatttca
     2281 agaatgcatg cgtcaatttt acgcagacta tctttctagg gttaaaaaag atttgcgctt
     2341 tactcgacct aaactttaaa cacgtcatag aatcttcgtt tgacaaaaac cacattgtgg
     2401 ggtaccgagc tcgaattcat cgatgatatc agatctgccg gtctccctat agtgagtcgt
     2461 attaatttcg ataagccagg ttaacctgca ttaatgaatc ggccaacgcg cggggagagg
     2521 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     2581 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     2641 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     2701 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     2761 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     2821 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     2881 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     2941 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     3001 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     3061 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     3121 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     3181 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     3241 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     3301 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     3361 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     3421 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     3481 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     3541 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     3601 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     3661 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     3721 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     3781 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     3841 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     3901 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     3961 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     4021 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     4081 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     4141 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     4201 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     4261 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     4321 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     4381 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     4441 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
     4501 attaacctat aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga
     4561 cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga
     4621 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg
     4681 gcttaactat gcggcatcag agcagattgt actgagagtg caccatatgg acatattgtc
     4741 gttagaacgc ggctacaatt aatacataac cttatgtatc atacacatac gatttaggtg
     4801 acactataga actcgacctc gaggctggca cgacaggttt cccgactgga aagcgggcag
     4861 tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt
     4921 tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa
     4981 cagctatgac catgattacg ccaagctcga aattaaccct cactaaaggg aacaaaagct
     5041 ggagctcgtc tttgatcaaa acgcaaatcg acgaaaatgt gtcggacaat atcaagtcga
     5101 tgagcgaaaa actaaaaagg ctagaatacg acaatctcac agacagcgtt gagatatacg
     5161 gtattcacga cagcaggctg aataataaaa aaattagaaa ctattattta accctagaaa
     5221 gataatcata ttgtgacgta cgttaaagat aatcatgcgt aaaattgacg catgtgtttt
     5281 atcgatctgt atatcgaggt ttatttatta atttgaatag atattaagtt ttattatatt
     5341 tacacttaca tactaataat aaattcaaca aacaatttat ttatgtttat ttatttatta
     5401 aaaaaaaaca aaaactcaaa atttcttcta taaagtaaca aaacttttaa acattctctc
     5461 ttttacaaaa ataaacttat tttgtacttt aaaaacagtc atgttgtatt ataaaataag
     5521 taattagctt aacttataca taatagaaac aaattatact tattagacat tgattattga
     5581 ctagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
     5641 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
     5701 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
     5761 aatgggtgga
//
