LOCUS       sequence-129058-        5937 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid encoding perch Cx35 with a C-terminal 
            HaloTag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5937)
  AUTHORS   Wang HY, Lin YP, Mitchell CK, Ram S, O'Brien J
  TITLE     Two-color fluorescent analysis of connexin 36 turnover: relationship
            to functional plasticity.
  JOURNAL   J Cell Sci. 2015 Nov 1;128(21):3888-97. doi: 10.1242/jcs.162586. 
            Epub 2015 Sep 10.
  PUBMED    26359298
REFERENCE   2  (bases 1 to 5937)
  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: 
            "10.1242/jcs.162586"; journalName: "J Cell Sci"; date: "2015-11-1- 
            1"; volume: "128"; issue: "21"; pages: "3888-97"
COMMENT     Sequence Label: CMV-Cx35-Halo-C-end
FEATURES             Location/Qualifiers
     source          1..5937
                     /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"
     CDS             1580..1600
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             1616..2500
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     primer_bind     complement(2572..2590)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2725..2846
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2762..2781)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2816..2835
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2853..3308)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2990..3011)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3202..3221
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3335..3439
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3441..3798
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3441..3632
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3464..3484)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3649..3784
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3833..4627
                     /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             3833..4627
                     /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(3887..3906)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4497..4516
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4815..4834)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4859..4906
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5235..5823
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5724..5743
                     /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 atctcgagct caagcttcga attctccgga tgctcagaaa tgggggaatg
      661 gactatacta gagaggctcc tggaggctgc tgtccagcag cactctacta tgataggaag
      721 gatcctacta acagtggtgg tcatcttccg gattctaatc gtagcaatag ttggagagac
      781 tgtctatgat gacgagcaga ccatgtttgt ttgtaacacc ttgcaaccgg gctgcaacca
      841 ggcatgctac gacaaggcat ttcccatttc acacataaga tattgggttt ttcagatcat
      901 catggtgtgc accccgagtc tttgttttat aacgtactcg gtgcaccagt cggccaagca
      961 gaaggagcgg cggtactcaa cagtctatct gacactggat aaggatcaag attcactgaa
     1021 gcgagacgag agcaaaaaga taaagaacac cattgttaat ggagtacttc agaacacgga
     1081 gaactccacc aaagaagccg agccagactg tttagaagtc aaagagatcc ctaattcggc
     1141 catgagaact acaaagtcca aaatgaggcg acaagagggc atctccaggt tttacatcat
     1201 ccaggtggtt ttcagaaacg cgctggaaat aggcttcttg gtgggtcagt atttcttgta
     1261 tggattcaac gtcccgtcgg tgtacgagtg cgatcgctac ccctgcatca aagatgtcga
     1321 gtgctacgtc tccagaccca cggagaagac agtgttcctg gtcttcatgt tcgcggtcag
     1381 cgggttctgc gtggtgctga acctggcaga actcaatcac ctgggctgga ggaaaatcaa
     1441 aacggccgtg agaggagtgc aggctcggcg gaagtccatt tatgagatca gaaataagga
     1501 cttgccgagg atgagtgtgc ctaatttcgg ccgtactcag tccagtgact ctgcttatgt
     1561 gtccctcgag ccaaccactg aggatctgta ctttcagagc gataacgatg gatccgaaat
     1621 cggtactggc tttccattcg acccccatta tgtggaagtc ctgggcgagc gcatgcacta
     1681 cgtcgatgtt ggtccgcgcg atggcacccc tgtgctgttc ctgcacggta acccgacctc
     1741 ctcctacgtg tggcgcaaca tcatcccgca tgttgcaccg acccatcgct gcattgctcc
     1801 agacctgatc ggtatgggca aatccgacaa accagacctg ggttatttct tcgacgacca
     1861 cgtccgcttc atggatgcct tcatcgaagc cctgggtctg gaagaggtcg tcctggtcat
     1921 tcacgactgg ggctccgctc tgggtttcca ctgggccaag cgcaatccag agcgcgtcaa
     1981 aggtattgca tttatggagt tcatccgccc tatcccgacc tgggacgaat ggccagaatt
     2041 tgcccgcgag accttccagg ccttccgcac caccgacgtc ggccgcaagc tgatcatcga
     2101 tcagaacgtt tttatcgagg gtacgctgcc gatgggtgtc gtccgcccgc tgactgaagt
     2161 cgagatggac cattaccgcg agccgttcct gaatcctgtt gaccgcgagc cactgtggcg
     2221 cttcccaaac gagctgccaa tcgccggtga gccagcgaac atcgtcgcgc tggtcgaaga
     2281 atacatggac tggctgcacc agtcccctgt cccgaagctg ctgttctggg gcaccccagg
     2341 cgttctgatc ccaccggccg aagccgctcg cctggccaaa agcctgccta actgcaaggc
     2401 tgtggacatc ggcccgggtc tgaatctgct gcaagaagac aacccggacc tgatcggcag
     2461 cgagatcgcg cgctggctgt ctactctgga gatttccggt taatagaatt ggcatgcaag
     2521 ctgatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc accgctgagc
     2581 aataactagc ataacccctt ggggcggccg cgactctaga tcataatcag ccataccaca
     2641 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
     2701 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
     2761 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     2821 ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta agcgttaata ttttgttaaa
     2881 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     2941 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     3001 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     3061 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     3121 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     3181 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     3241 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     3301 gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     3361 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     3421 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
     3481 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3541 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3601 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     3661 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     3721 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     3781 ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga
     3841 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     3901 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     3961 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga
     4021 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     4081 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     4141 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     4201 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     4261 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     4321 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga
     4381 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     4441 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     4501 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     4561 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     4621 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     4681 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     4741 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct
     4801 agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg
     4861 gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt
     4921 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc
     4981 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca
     5041 gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt
     5101 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     5161 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     5221 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     5281 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     5341 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     5401 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     5461 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     5521 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     5581 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     5641 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     5701 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     5761 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     5821 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     5881 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcat
//
