LOCUS       sequence-132221-        7031 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid encoding perch Cx35 with an internal 
            HaloTag between amino acids 285 and 286 in the C-terminus..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7031)
  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 7031)
  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-in
FEATURES             Location/Qualifiers
     source          1..7031
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1949..2839
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             2852..2872
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     primer_bind     complement(3038..3055)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3044..3268
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3314..3742
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3401..3420)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3611..3632
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3751..3771)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3756..4086
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3937..4072
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        4134..4181
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4200..4574
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    4704..4825
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4741..4760)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4795..4814
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4874..4890)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4874..4890)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4887..4909)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4898..4914
                     /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(4922..4928,4929..4946,4947..4952))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4967..4988
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5105..5122)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5276..5864)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5356..5375)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6035..6826,6827..6895))
                     /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     6658..6677
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6896..7000)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattctgc
      961 agatctaaaa tcggacgctt tggagactga gaacaacgag aggcagggag aagaaggacg
     1021 gggaaataac gcgctttttt ggagggattg aacagagtgc cgttgtgctc tagtcttctc
     1081 cggatgctca gaaatggggg aatggactat actagagagg ctcctggagg ctgctgtcca
     1141 gcagcactct actatgatag gaaggatcct actaacagtg gtggtcatct tccggattct
     1201 aatcgtagca atagttggag agactgtcta tgatgacgag cagaccatgt ttgtttgtaa
     1261 caccttgcaa ccgggctgca accaggcatg ctacgacaag gcatttccca tttcacacat
     1321 aagatattgg gtttttcaga tcatcatggt gtgcaccccg agtctttgtt ttataacgta
     1381 ctcggtgcac cagtcggcca agcagaagga gcggcggtac tcaacagtct atctgacact
     1441 ggataaggat caagattcac tgaagcgaga cgagagcaaa aagataaaga acaccattgt
     1501 taatggagta cttcagaaca cggagaactc caccaaagaa gccgagccag actgtttaga
     1561 agtcaaagag atccctaatt cggccatgag aactacaaag tccaaaatga ggcgacaaga
     1621 gggcatctcc aggttttaca tcatccaggt ggttttcaga aacgcgctgg aaataggctt
     1681 cttggtgggt cagtatttct tgtatggatt caacgtcccg tcggtgtacg agtgcgatcg
     1741 ctacccctgc atcaaagatg tcgagtgcta cgtctccaga cccacggaga agacagtgtt
     1801 cctggtcttc atgttcgcgg tcagcgggtt ctgcgtggtg ctgaacctgg cagaactcaa
     1861 tcacctgggc tggaggaaaa tcaaaacggc cgtgagagga gtgcaggctc ggcggaagtc
     1921 catttatgag atcagaaata aggacttgat ggcagaaatc ggtactggct ttccattcga
     1981 cccccattat gtggaagtcc tgggcgagcg catgcactac gtcgatgttg gtccgcgcga
     2041 tggcacccct gtgctgttcc tgcacggtaa cccgacctcc tcctacgtgt ggcgcaacat
     2101 catcccgcat gttgcaccga cccatcgctg cattgctcca gacctgatcg gtatgggcaa
     2161 atccgacaaa ccagacctgg gttatttctt cgacgaccac gtccgcttca tggatgcctt
     2221 catcgaagcc ctgggtctgg aagaggtcgt cctggtcatt cacgactggg gctccgctct
     2281 gggtttccac tgggccaagc gcaatccaga gcgcgtcaaa ggtattgcat ttatggagtt
     2341 catccgccct atcccgacct gggacgaatg gccagaattt gcccgcgaga ccttccaggc
     2401 cttccgcacc accgacgtcg gccgcaagct gatcatcgat cagaacgttt ttatcgaggg
     2461 tacgctgccg atgggtgtcg tccgcccgct gactgaagtc gagatggacc attaccgcga
     2521 gccgttcctg aatcctgttg accgcgagcc actgtggcgc ttcccaaacg agctgccaat
     2581 cgccggtgag ccagcgaaca tcgtcgcgct ggtcgaagaa tacatggact ggctgcacca
     2641 gtcccctgtc ccgaagctgc tgttctgggg caccccaggc gttctgatcc caccggccga
     2701 agccgctcgc ctggccaaaa gcctgcctaa ctgcaaggct gtggacatcg gcccgggtct
     2761 gaatctgctg caagaagaca acccggacct gatcggcagc gagatcgcgc gctggctgtc
     2821 gacgctcgag atttccggcg agccaaccac tgaggatctg tactttcaga gcccgaggat
     2881 gagtgtgcct aatttcggcc gtactcagtc cagtgactct gcttatgtgt aacaaacaca
     2941 cacatacaca cacacaaacg gggaggaaga agtgaaaata tgtaaaaaaa aaaaaaaaaa
     3001 actcgagtct agagggcccg tttaaaccgc tgatcagcct cgactgtgcc ttctagttgc
     3061 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     3121 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     3181 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     3241 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     3301 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     3361 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     3421 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg catcccttta
     3481 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     3541 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     3601 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     3661 tcttttgatt tataagggat tttggggatt tcggcctatt ggttaaaaaa tgagctgatt
     3721 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt
     3781 ccccaggctc cccaggcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     3841 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3901 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     3961 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     4021 gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     4081 tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcagca cgtgttgaca
     4141 attaatcatc ggcatagtat atcggcatag tataatacga caaggtgagg aactaaacca
     4201 tggccaagtt gaccagtgcc gttccggtgc tcaccgcgcg cgacgtcgcc ggagcggtcg
     4261 agttctggac cgaccggctc gggttctccc gggacttcgt ggaggacgac ttcgccggtg
     4321 tggtccggga cgacgtgacc ctgttcatca gcgcggtcca ggaccaggtg gtgccggaca
     4381 acaccctggc ctgggtgtgg gtgcgcggcc tggacgagct gtacgccgag tggtcggagg
     4441 tcgtgtccac gaacttccgg gacgcctccg ggccggccat gaccgagatc ggcgagcagc
     4501 cgtgggggcg ggagttcgcc ctgcgcgacc cggccggcaa ctgcgtgcac ttcgtggccg
     4561 aggagcagga ctgacacgtg ctacgagatt tcgattccac cgccgccttc tatgaaaggt
     4621 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     4681 tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa
     4741 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     4801 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct
     4861 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     4921 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     4981 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     5041 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     5101 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     5161 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     5221 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     5281 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     5341 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     5401 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     5461 cgctttctca atgctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5521 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5581 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5641 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5701 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5761 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5821 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5881 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5941 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     6001 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     6061 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     6121 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     6181 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     6241 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6301 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     6361 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6421 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6481 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6541 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6601 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     6661 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6721 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     6781 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     6841 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     6901 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6961 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     7021 cacctgacgt c
//
