LOCUS       sequence-199360-        8793 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To visualize the expression and localization of potassium-chloride 
            cotransporter type 2 (KCC2).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8793)
  AUTHORS   Friedel P, Ludwig A, Pellegrino C, Agez M, Jawhari A, Rivera C, 
            Medina I
  TITLE     A Novel View on the Role of Intracellular Tails in Surface Delivery 
            of the Potassium-Chloride Cotransporter KCC2.
  JOURNAL   eNeuro. 2017 Jul 21;4(4). pii: ENEURO.0055-17.2017. doi: 
            10.1523/ENEURO.0055-17.2017. eCollection 2017 Jul-Aug.
  PUBMED    28785725
REFERENCE   2  (bases 1 to 8793)
  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: "eNeuro."; 
            date: "2017-07-21"; volume: "4(4). pii: ENEURO.0055-17.2017. doi"; 
            pages: " 10.1523/ENEURO.0055-17.2017. eCollection 2017 Jul-Aug"
COMMENT     Sequence Label: KCC2-pHext
FEATURES             Location/Qualifiers
     source          1..8793
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..1212
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     1049..1068
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             2009..2716
                     /codon_start=1
                     /product="pH-sensitive mutant of green fluorescent protein 
                     (Ng et al., 2002)"
                     /label=superecliptic pHluorin
                     /note="excited strongly by blue light at neutral pH but 
                     minimally fluorescent at acidic pH"
                     /translation="KGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFI
                     CTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGNY
                     KTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNDHQVYIMADKQKNGIKANF
                     KIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLFTTSTLSKDPNEKRDHMVLLEFV
                     TAAGITHGMDELYK"
     primer_bind     complement(2033..2061)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2656..2676
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             complement(2882..2893)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    5563..5684
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5600..5619)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5654..5673
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5691..6146)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5828..5849)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6040..6059
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6173..6277
                     /gene="bla"
                     /label=AmpR promoter
     promoter        6279..6636
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6279..6470
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6302..6322)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6487..6622
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6671..7465
                     /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             6671..7465
                     /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(6725..6744)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7335..7354
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7653..7672)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7697..7744
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      8073..8661
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8562..8581
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc cctcctcacg gcgagcgctg
       61 ccacgtcaga cgaagggcgc agcgagcgtc ctgatccttc cgcccggacg ctcaggacag
      121 cggcccgctg ctcataagac tcggccttag aaccccagta tcagcagaag gacattttag
      181 gacgggactt gggtgactct agggcactgg ttttctttcc agagagcgga acaggcgagg
      241 aaaagtagtc ccttctcggc gattctgcgg agggatctcc gtggggcggt gaacgccgat
      301 gattatataa ggacgcgccg ggtgtggcac agctagttcc gtcgcagccg ggatttgggt
      361 cgcggttctt gtttgtggat cgctgtgatc gtcacttggt gagtagcggg ctgctgggct
      421 ggccggggct ttcgtggccg ccgggccgct cggtgggacg gaagcgtgtg gagagaccgc
      481 caagggctgt agtctgggtc cgcgagcaag gttgccctga actgggggtt ggggggagcg
      541 cagcaaaatg gcggctgttc ccgagtcttg aatggaagac gcttgtgagg cgggctgtga
      601 ggtcgttgaa acaaggtggg gggcatggtg ggcggcaaga acccaaggtc ttgaggcctt
      661 cgctaatgcg ggaaagctct tattcgggtg agatgggctg gggcaccatc tggggaccct
      721 gacgtgaagt ttgtcactga ctggagaact cggtttgtcg tctgttgcgg gggcggcagt
      781 tatggcggtg ccgttgggca gtgcacccgt acctttggga gcgcgcgccc tcgtcgtgtc
      841 gtgacgtcac ccgttctgtt ggcttataat gcagggtggg gccacctgcc ggtaggtgtg
      901 cggtaggctt ttctccgtcg caggacgcag ggttcgggcc tagggtaggc tctcctgaat
      961 cgacaggcgc cggacctctg gtgaggggag ggataagtga ggcgtcagtt tctttggtcg
     1021 gttttatgta cctatcttct taagtagctg aagctccggt tttgaactat gcgctcgggg
     1081 ttggcgagtg tgttttgtga agttttttag gcaccttttg aaatgtaatc atttgggtca
     1141 atatgtaatt ttcagtgtta gactagtaaa ttgtccgcta aattctggcc gtttttggct
     1201 tttttgttag acgaagcttc aagcttcggc acgaggcgaa ggcgggcaga ggggcgcggg
     1261 cgaagaggcg cagccatccc gagcccggcg ccgcgcagcc accatgctca acaacctgac
     1321 ggactgcgag gacggcgatg ggggagccaa cccgggtgac ggcaatccca aggagagcag
     1381 ccccttcatc aacagcacgg acacggagaa ggggagagag tatgatggca ggaacatggc
     1441 cctgtttgag gaggagatgg acaccagccc catggtatcc tccctgctca gtgggctggc
     1501 caactacacc aacctgcctc agggaagcaa agagcacgaa gaagcagaaa acaatgaggg
     1561 cggaaagaag aagccggtgc aggccccacg catgggcacc ttcatgggcg tgtacctccc
     1621 gtgcctgcag aacatctttg gtgttatcct ctttctgcgg ctcacttggg tggtgggaat
     1681 cgcaggcatc atggagtcct tctgcatggt cttcatctgc tgctcctgca cgatgctcac
     1741 agccatttcc atgagcgcaa ttgcaaccaa tggtgttgtg cctgctggtg gctcctacta
     1801 catgatttcc aggtctctgg gcccggagtt tgggggcgcc gtgggcctct gcttctacct
     1861 gggcactacc tttgctgggg ctatgtacat cctgggcacc atcgagatcc tgctggctta
     1921 cctcttccca gcgatggcca tcttcaaggc agaagatgcc agtggggagg cagccgccat
     1981 gattaatgaa gaagacctag gcggccgcaa aggagaagaa cttttcactg gagttgtccc
     2041 aattcttgtt gaattagatg gtgatgttaa tgggcacaaa ttttctgtca gtggagaggg
     2101 tgaaggtgat gcaacatacg gaaaacttac ccttaaattt atttgcacta ctggaaaact
     2161 acctgttcca tggccaacac ttgtcactac tctgacctat ggtgttcaat gcttttcaag
     2221 atacccagat catatgaaac ggcatgactt tttcaagagt gccatgcccg aaggttatgt
     2281 acaggaaaga actatatttt tcaaagatga cgggaactac aagacacgtg ctgaagtcaa
     2341 gtttgaaggt gatacccttg ttaatagaat cgagttaaaa ggtattgatt ttaaagaaga
     2401 tggaaacatt cttggacaca aattggaata caactataac gatcaccagg tgtacatcat
     2461 ggcagacaaa caaaagaatg gaatcaaagc taacttcaaa attagacaca acattgaaga
     2521 tggaggcgtt caactagcag accattatca acaaaatact ccaattggcg atgggcccgt
     2581 ccttttacca gacaaccatt acctgtttac aacttctact ctttcgaaag atcccaacga
     2641 aaagagagac cacatggtcc ttcttgagtt tgtaacagct gctgggatta cacatggcat
     2701 ggatgaacta tacaaactcg aggatgaact tattaataac atgcgggtgt atggcacctg
     2761 tgtgctcacc tgcatggcca ccgtagtctt tgtgggcgtc aagtacgtga acaagtttgc
     2821 cctggtcttc ctgggttgcg tgatcctctc catcctggcc atctacgcag gggtcatcaa
     2881 gtctgccttc gatccaccca atttcccgat ttgcctcctg gggaaccgca cgctgtctcg
     2941 ccatggcttt gatgtctgtg ccaagctggc ttgggaagga aatgagacag tgaccacacg
     3001 gctctggggc ctattctgtt cctcccgcct cctcaatgcc acctgtgatg agtacttcac
     3061 ccgaaacaat gtcacagaga tccagggcat tcctggtgct gcaagtggcc tcatcaaaga
     3121 gaacctgtgg agttcctacc tgaccaaggg ggtgatcgtg gagaggcgtg ggatgccctc
     3181 tgtgggcctg gcagatggta cccccgttga catggaccac ccctatgtct tcagtgatat
     3241 gacctcctac ttcaccctgc ttgttggcat ctatttcccc tcagtcacag ggatcatggc
     3301 tggctcgaac cggtccggag acctgcggga tgcccagaag tctatcccta ctggaactat
     3361 cttggccatt gctacgacct ctgctgtcta catcagctct gttgttctgt tcggagcctg
     3421 catcgaaggg gtcgtcctac gggacaagtt tggggaagct gtcaacggga atctggtggt
     3481 gggcaccctg gcctggcctt ctccttgggt cattgtcata ggctctttct tctctacctg
     3541 cggagctgga ctacagagcc tcacaggggc cccacgcctg ctgcaggcca tctcccggga
     3601 tggcatagtg cccttcctgc aggtctttgg ccatggcaaa gccaacggag agccaacctg
     3661 ggcgctgctg ctgactgcct gcatctgtga gatcggcatc ctcatcgcct ccctggatga
     3721 ggtcgcccct atcctttcca tgttcttcct gatgtgttac atgtttgtga acttggcttg
     3781 cgcggtgcag acactgctga ggacgcccaa ctggaggcca cgcttccgat attaccactg
     3841 gaccctctcc ttcctgggca tgagcctctg cctggccctg atgttcattt gctcctggta
     3901 ttatgcgctg gtagctatgc tcatcgctgg cctcatctat aagtacatcg agtaccgggg
     3961 ggcagagaag gagtgggggg atgggatccg aggcctgtct ctcagtgcag ctcgctatgc
     4021 tctcttgcgt ctggaggaag gacccccgca tacaaagaac tggaggcccc agctactggt
     4081 gctggtgcgt gtggaccagg accagaacgt ggtgcacccg cagctgctgt ccttgacctc
     4141 ccagctcaag gcagggaagg gcctgaccat tgtgggctct gtccttgagg gcacctttct
     4201 ggacaaccac cctcaggctc agcgggcaga ggagtctatc cggcgcctga tggaggctga
     4261 gaaggtgaag ggcttctgcc aggtagtgat ctcctccaac ctgcgtgacg gtgtgtccca
     4321 cctgatccaa tccgggggcc tcgggggcct gcaacacaac actgtgctag tgggctggcc
     4381 tcgcaactgg cgacagaagg aggatcatca gacatggagg aacttcatcg aactcgtccg
     4441 ggaaactaca gctggccacc tcgccctgct ggtcaccaag aatgtttcca tgttccccgg
     4501 gaaccctgag cgtttctctg agggcagcat tgacgtgtgg tggatcgtgc acgacggggg
     4561 catgctcatg ctgttgccct tcctcctgcg tcaccacaag gtctggagga aatgcaaaat
     4621 gcggatcttc accgtggcgc agatggatga caacagcatt cagatgaaga aagacctgac
     4681 cacgtttctg taccacttac gaattactgc agaggtggaa gtcgtggaga tgcacgagag
     4741 cgacatctca gcatacacct acgagaagac attagtcatg gagcaacgtt ctcagatcct
     4801 caaacagatg cacctcacca agaacgagcg ggaacgggag atccagagca tcacagatga
     4861 atctcggggc tccattcgga ggaagaatcc agccaacact cggctccgcc tcaatgttcc
     4921 cgaagagaca gcttgtgaca acgaggagaa gccagaagag gaggtgcagc tgatccatga
     4981 ccagagtgct cccagctgcc ctagcagctc gccgtctcca ggggaggagc ctgaggggga
     5041 gggggagaca gacccagaga aggtgcatct cacctggacc aaggataagt cagcggctca
     5101 gaagaacaaa ggccccagtc ccgtctcctc ggaggggatc aaggacttct tcagcatgaa
     5161 gccggagtgg gaaaacttga accagtccaa cgtgcggcgc atgcacacag ctgtgcggct
     5221 gaacgaggtc atcgtgaata aatcccggga tgccaagttg gtgttgctca acatgcccgg
     5281 gcctccccgc aaccgcaatg gagatgaaaa ctacatggag ttcctggagg tcctcactga
     5341 gcaactggac cgggtgatgc tggtccgcgg tggtggccga gaggtcatca ccatctactc
     5401 ctgaattctg cagtcgacgg taccgcgggc ccgggatcca ccggatctag ataactgatc
     5461 ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc
     5521 cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct
     5581 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     5641 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaacgcgt aaattgtaag
     5701 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     5761 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     5821 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     5881 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     5941 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     6001 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     6061 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     6121 gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac
     6181 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     6241 ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg
     6301 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     6361 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     6421 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     6481 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     6541 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     6601 tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg
     6661 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     6721 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     6781 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     6841 gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     6901 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     6961 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     7021 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     7081 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     7141 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag
     7201 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     7261 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     7321 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     7381 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     7441 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     7501 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     7561 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     7621 gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa
     7681 ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt
     7741 gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc
     7801 cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt
     7861 gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac
     7921 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     7981 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     8041 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     8101 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     8161 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     8221 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     8281 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     8341 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     8401 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     8461 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     8521 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     8581 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     8641 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     8701 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     8761 attaccgcca tgcattagtt attaccggga tct
//
