LOCUS       sequence-199357-        8821 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 8821)
  AUTHORS   Kahle KT, Merner ND, Friedel P, Silayeva L, Liang B, Khanna A, Shang
            Y, Lachance-Touchette P, Bourassa C, Levert A, Dion PA, Walcott B, 
            Spiegelman D, Dionne-Laporte A, Hodgkinson A, Awadalla P, Nikbakht 
            H, Majewski J, Cossette P, Deeb TZ, Moss SJ, Medina I, Rouleau GA
  TITLE     Genetically encoded impairment of neuronal KCC2 cotransporter 
            function in human idiopathic generalized epilepsy.
  JOURNAL   EMBO Rep. 2014 Jul;15(7):766-74. doi: 10.15252/embr.201438840. Epub 
            2014 Jun 13.
  PUBMED    24928908
REFERENCE   2  (bases 1 to 8821)
  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: 
            "10.15252/embr.201438840"; journalName: "EMBO Rep"; date: "2014-07";
            volume: "15"; issue: "7"; pages: "766-74"
COMMENT     Sequence Label: Cherry-HA-KCC2
FEATURES             Location/Qualifiers
     source          1..8821
                     /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             1239..1946
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1384..1402)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1652..1671
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1962..1988
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2910..2921)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    5591..5712
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5628..5647)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5682..5701
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5719..6174)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5856..5877)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6068..6087
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6201..6305
                     /gene="bla"
                     /label=AmpR promoter
     promoter        6307..6664
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6307..6498
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6330..6350)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6515..6650
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6699..7493
                     /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             6699..7493
                     /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(6753..6772)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7363..7382
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7681..7700)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7725..7772
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      8101..8689
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8590..8609
                     /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 acgaagcttg ctagcgctaa gatctgccat ggtgagcaag ggcgaggagg
     1261 ataacatggc catcatcaag gagttcatgc gcttcaaggt gcacatggag ggctccgtga
     1321 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
     1381 ccgccaagct gaaggtgacc aagggtggcc ccctgccctt cgcctgggac atcctgtccc
     1441 ctcagttcat gtacggctcc aaggcctacg tgaagcaccc cgccgacatc cccgactact
     1501 tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
     1561 gcgtggtgac cgtgacccag gactcctccc tgcaggacgg cgagttcatc tacaaggtga
     1621 agctgcgcgg caccaacttc ccctccgacg gccccgtaat gcagaagaag accatgggct
     1681 gggaggcctc ctccgagcgg atgtaccccg aggacggcgc cctgaagggc gagatcaagc
     1741 agaggctgaa gctgaaggac ggcggccact acgacgctga ggtcaagacc acctacaagg
     1801 ccaagaagcc cgtgcagctg cccggcgcct acaacgtcaa catcaagttg gacatcacct
     1861 cccacaacga ggactacacc atcgtggaac agtacgaacg cgccgagggc cgccactcca
     1921 ccggcggcat ggacgagctg tacaagctca gatctgccat gtacccatac gatgttccag
     1981 attacgcttc tcgagctcaa gcttcggcac gaggcgaagg cgggcagagg ggcgcgggcg
     2041 aagaggcgca gccatcccga gcccggcgcc gcgcagccac catgctcaac aacctgacgg
     2101 actgcgagga cggcgatggg ggagccaacc cgggtgacgg caatcccaag gagagcagcc
     2161 ccttcatcaa cagcacggac acggagaagg ggagagagta tgatggcagg aacatggccc
     2221 tgtttgagga ggagatggac accagcccca tggtatcctc cctgctcagt gggctggcca
     2281 actacaccaa cctgcctcag ggaagcaaag agcacgaaga agcagaaaac aatgagggcg
     2341 gaaagaagaa gccggtgcag gccccacgca tgggcacctt catgggcgtg tacctcccgt
     2401 gcctgcagaa catctttggt gttatcctct ttctgcggct cacttgggtg gtgggaatcg
     2461 caggcatcat ggagtccttc tgcatggtct tcatctgctg ctcctgcacg atgctcacag
     2521 ccatttccat gagcgcaatt gcaaccaatg gtgttgtgcc tgctggtggc tcctactaca
     2581 tgatttccag gtctctgggc ccggagtttg ggggcgccgt gggcctctgc ttctacctgg
     2641 gcactacctt tgctggggct atgtacatcc tgggcaccat cgagatcctg ctggcttacc
     2701 tcttcccagc gatggccatc ttcaaggcag aagatgccag tggggaggca gccgccatgt
     2761 tgaataacat gcgggtgtat ggcacctgtg tgctcacctg catggccacc gtagtctttg
     2821 tgggcgtcaa gtacgtgaac aagtttgccc tggtcttcct gggttgcgtg atcctctcca
     2881 tcctggccat ctacgcaggg gtcatcaagt ctgccttcga tccacccaat ttcccgattt
     2941 gcctcctggg gaaccgcacg ctgtctcgcc atggctttga tgtctgtgcc aagctggctt
     3001 gggaaggaaa tgagacagtg accacacggc tctggggcct attctgttcc tcccgcctcc
     3061 tcaatgccac ctgtgatgag tacttcaccc gaaacaatgt cacagagatc cagggcattc
     3121 ctggtgctgc aagtggcctc atcaaagaga acctgtggag ttcctacctg accaaggggg
     3181 tgatcgtgga gaggcgtggg atgccctctg tgggcctggc agatggtacc cccgttgaca
     3241 tggaccaccc ctatgtcttc agtgatatga cctcctactt caccctgctt gttggcatct
     3301 atttcccctc agtcacaggg atcatggctg gctcgaaccg gtccggagac ctgcgggatg
     3361 cccagaagtc tatccctact ggaactatct tggccattgc tacgacctct gctgtctaca
     3421 tcagctctgt tgttctgttc ggagcctgca tcgaaggggt cgtcctacgg gacaagtttg
     3481 gggaagctgt gaatggcaat ctggtggtgg gcaccctggc ctggccttct ccttgggtca
     3541 ttgtcatagg ctctttcttc tctacctgcg gagctggact acagagcctc acaggggccc
     3601 cacgcctgct gcaggccatc tcccgggatg gcatagtgcc cttcctgcag gtctttggcc
     3661 atggcaaagc caacggagag ccaacctggg cgctgctgct gactgcctgc atctgtgaga
     3721 tcggcatcct catcgcctcc ctggatgagg tcgcccctat cctttccatg ttcttcctga
     3781 tgtgttacat gtttgtgaac ttggcttgcg cggtgcagac actgctgagg acgcccaact
     3841 ggaggccacg cttccgatat taccactgga ccctctcctt cctgggcatg agcctctgcc
     3901 tggccctgat gttcatttgc tcctggtatt atgcgctggt agctatgctc atcgctggcc
     3961 tcatctataa gtacatcgag taccgggggg cagagaagga gtggggggat gggatccgag
     4021 gcctgtctct cagtgcagct cgctatgctc tcttgcgtct ggaggaagga cccccgcata
     4081 caaagaactg gaggccccag ctactggtgc tggtgcgtgt ggaccaggac cagaacgtgg
     4141 tgcacccgca gctgctgtcc ttgacctccc agctcaaggc agggaagggc ctgaccattg
     4201 tgggctctgt ccttgagggc acctttctgg acaaccaccc tcaggctcag cgggcagagg
     4261 agtctatccg gcgcctgatg gaggctgaga aggtgaaggg cttctgccag gtagtgatct
     4321 cctccaacct gcgtgacggt gtgtcccacc tgatccaatc cgggggcctc gggggcctgc
     4381 aacacaacac tgtgctagtg ggctggcctc gcaactggcg acagaaggag gatcatcaga
     4441 catggaggaa cttcatcgaa ctcgtccggg aaactacagc tggccacctc gccctgctgg
     4501 tcaccaagaa tgtttccatg ttccccggga accctgagcg tttctctgag ggcagcattg
     4561 acgtgtggtg gatcgtgcac gacgggggca tgctcatgct gttgcccttc ctcctgcgtc
     4621 accacaaggt ctggaggaaa tgcaaaatgc ggatcttcac cgtggcgcag atggatgaca
     4681 acagcattca gatgaagaaa gacctgacca cgtttctgta ccacttacga attactgcag
     4741 aggtggaagt cgtggagatg cacgagagcg acatctcagc atacacctac gagaagacat
     4801 tggtaatgga acaacgttct cagatcctca aacagatgca cctcaccaag aacgagcggg
     4861 aacgggagat ccagagcatc acagatgaat ctcggggctc cattcggagg aagaatccag
     4921 ccaacactcg gctccgcctc aatgttcccg aagagacagc ttgtgacaac gaggagaagc
     4981 cagaagagga ggtgcagctg atccatgacc agagtgctcc cagctgccct agcagctcgc
     5041 cgtctccagg ggaggagcct gagggggagg gggagacaga cccagagaag gtgcatctca
     5101 cctggaccaa ggataagtca gcggctcaga agaacaaagg ccccagtccc gtctcctcgg
     5161 aggggatcaa ggacttcttc agcatgaagc cggagtggga aaacttgaac cagtccaacg
     5221 tgcggcgcat gcacacagct gtgcggctga acgaggtcat cgtgaataaa tcccgggatg
     5281 ccaagttggt gttgctcaac atgcccgggc ctccccgcaa ccgcaatgga gatgaaaact
     5341 acatggagtt cctggaggtc ctcactgagc aactggaccg ggtgatgctg gtccgcggtg
     5401 gtggccgaga ggtcatcacc atctactcct gaattctgca gtcgacggta ccgcgggccc
     5461 gggatccacc ggatctagat aactgatcat aatcagccat accacatttg tagaggtttt
     5521 acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat
     5581 tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     5641 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     5701 caatgtatct taacgcgtaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
     5761 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     5821 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     5881 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     5941 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     6001 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     6061 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     6121 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg
     6181 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     6241 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     6301 agagtcctga ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc
     6361 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     6421 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     6481 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     6541 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     6601 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     6661 caaagatcga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     6721 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     6781 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     6841 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat
     6901 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     6961 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     7021 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     7081 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     7141 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     7201 ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc
     7261 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     7321 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     7381 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     7441 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     7501 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     7561 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     7621 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac
     7681 tgaaacacgg aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca
     7741 gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg
     7801 gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt
     7861 ttccccaccc caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc
     7921 ggcaggccct gccatagcct caggttactc atatatactt tagattgatt taaaacttca
     7981 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     8041 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     8101 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     8161 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     8221 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     8281 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     8341 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     8401 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     8461 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     8521 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     8581 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     8641 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     8701 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     8761 gttatcccct gattctgtgg ataaccgtat taccgccatg cattagttat taccgggatc
     8821 t
//
