LOCUS       sequence-124710-        5888 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses rat Cx43 (Gja1 CDS) with an 8 amino acid linker on the 
            N-terminus of the Connexin linking to Enhanced Blue Fluorescent 
            Protein 2 (FP fused to NT of connexin). CMV promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5888)
  AUTHORS   Stout RF Jr, Snapp EL, Spray DC
  TITLE     Connexin Type and Fluorescent Protein-fusion Tag Determine 
            Structural Stability of Gap Junction Plaques.
  JOURNAL   J Biol Chem. 2015 Aug 11. pii: jbc.M115.659979.
  PUBMED    26265468
REFERENCE   2  (bases 1 to 5888)
  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: "J Biol 
            Chem. 2015 Aug 11. pii: jbc.M115.659979."
COMMENT     Sequence Label: EBFP2-Cx43
FEATURES             Location/Qualifiers
     source          1..5888
                     /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      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced blue variant of GFP (Ai et al., 2007)"
                     /label=EBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNFNSHNIYIMAVKQKNGIK
                     VNFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLL
                     EFRTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     polyA_signal    2676..2797
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2713..2732)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2767..2786
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2804..3259)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2941..2962)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3153..3172
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3286..3390
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3392..3749
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3392..3583
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3415..3435)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3600..3735
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3784..4578
                     /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             3784..4578
                     /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(3838..3857)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4448..4467
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4766..4785)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4810..4857
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5186..5774
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5675..5694
                     /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 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgaggggcga gggcgagggc
      721 gatgccacca acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgagc cacggcgtgc agtgcttcgc ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcacc tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcgtcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaacttc aacagccaca acatctatat catggccgtc
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacgtgga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacagcc actacctgag cacccagtcc gtgctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttccgcacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct cagatgggtg actggagtgc cttggggaag
     1381 cttctggaca aggtccaagc ctactccacc gctggaggga aggtgtggct gtcagtgctc
     1441 ttcatattca gaatcctgct cctggggaca gctgttgagt cagcttgggg tgatgaacag
     1501 tctgcctttc gctgtaacac tcaacaacct ggctgcgaaa acgtctgcta tgacaagtcc
     1561 ttccccatct ctcacgtgcg cttctgggtc cttcagatca tattcgtgtc tgtgcccacc
     1621 ctcctgtact tggcccatgt gttctatgtg atgaggaagg aagagaagct aaacaagaaa
     1681 gaagaggagc tcaaagtggc ccagactgac ggggtcaacg tggagatgca cctgaagcag
     1741 attgaaatca agaagttcaa gtacgggatt gaagagcacg gcaaggtgaa aatgaggggc
     1801 ggcttgctga gaacctacat catcagcatc ctcttcaagt ctgtcttcga ggtggccttc
     1861 ctgctcatcc agtggtacat ctatgggttc agcttgagcg cggtctacac ctgcaagaga
     1921 gatccctgcc cccaccaggt agactgcttc ctctcacgtc ccacggagaa aaccatcttc
     1981 atcatcttca tgctggtggt gtccttggtg tctctcgctt tgaacatcat tgagctcttc
     2041 tacgtcttct tcaaaggcgt taaggatcgc gtgaagggaa gaagcgatcc ttaccacgcc
     2101 accactggcc cactgagccc atcaaaagac tgcggatctc caaaatacgc ctacttcaat
     2161 ggctgctcct caccaacggc tccactctcg cctatgtctc ctcctgggta caagctggtt
     2221 actggtgaca gaaacaattc ctcgtgccgc aattacaaca agcaagctag cgagcaaaac
     2281 tgggcgaact acagcgcaga gcaaaatcgc atggggcagg ccggaagcac catctccaac
     2341 tcgcacgccc agccgttcga tttccccgac gacaaccaga atgccaaaaa agttgctgct
     2401 ggacatgaac tccagccatt agccatcgtg gaccaacgac cttccagcag agccagcagc
     2461 cgcgccagca gcaggcctcg gcctgatgac ctggagattt aagagctcaa gcttcgaatt
     2521 ctgcagtcga cggtaccgcg ggcccgggat ccaccggatc tagataactg atcataatca
     2581 gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga
     2641 acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg
     2701 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     2761 ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg cgtaaattgt aagcgttaat
     2821 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc
     2881 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt
     2941 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa
     3001 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg
     3061 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga
     3121 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct
     3181 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat
     3241 gcgccgctac agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     3301 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     3361 atgcttcaat aatattgaaa aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg
     3421 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     3481 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     3541 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     3601 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     3661 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     3721 gcttttttgg aggcctaggc ttttgcaaag atcgatcaag agacaggatg aggatcgttt
     3781 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     3841 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     3901 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     3961 ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     4021 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     4081 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     4141 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     4201 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     4261 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc
     4321 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     4381 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     4441 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     4501 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     4561 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     4621 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     4681 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     4741 tcgcccaccc tagggggagg ctaactgaaa cacggaagga gacaataccg gaaggaaccc
     4801 gcgctatgac ggcaataaaa agacagaata aaacgcacgg tgttgggtcg tttgttcata
     4861 aacgcggggt tcggtcccag ggctggcact ctgtcgatac cccaccgaga ccccattggg
     4921 gccaatacgc ccgcgtttct tccttttccc caccccaccc cccaagttcg ggtgaaggcc
     4981 cagggctcgc agccaacgtc ggggcggcag gccctgccat agcctcaggt tactcatata
     5041 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     5101 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     5161 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     5221 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     5281 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     5341 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     5401 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     5461 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     5521 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     5581 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     5641 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     5701 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     5761 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     5821 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     5881 ccatgcat
//
