LOCUS       sequence-215139-        7704 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Electrophysiology in HEK cell.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7704)
  AUTHORS   Chang A, Abderemane-Ali F, Hura GL, Rossen ND, Gate RE, Minor DL Jr.
  TITLE     A Calmodulin C-Lobe Ca(2+)-Dependent Switch Governs Kv7 Channel 
            Function.
  JOURNAL   Neuron. 2018 Feb 21;97(4):836-852.e6. doi: 
            10.1016/j.neuron.2018.01.035. Epub 2018 Feb 8.
  PUBMED    29429937
REFERENCE   2  (bases 1 to 7704)
  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: 
            "10.1016/j.neuron.2018.01.035"; journalName: "Neuron"; date: 
            "2018-02-21- 21"; volume: "97"; issue: "4"; pages: "836-852.e6"
COMMENT     Sequence Label: Kv7.4 (1-645, Delta-368-523)/pcDNA3.1
FEATURES             Location/Qualifiers
     source          1..7704
                     /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             2435..2455
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             join(2456..2458,2459..3169)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2456..2458,2459..3169)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2498..2519)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2759..2778)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3106..3127
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3170..3193
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     promoter        complement(3257..3275)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3258..3275)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3295..3312)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3301..3525
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3571..3999
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3658..3677)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3868..3889
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4008..4028)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4013..4342
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4193..4328
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4409..5203
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4463..4482)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5073..5092
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5377..5498
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5414..5433)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5468..5487
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5547..5563)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5547..5563)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5560..5582)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5571..5587
                     /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(5595..5601,5602..5619,5620..5625))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5640..5661
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5778..5795)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5949..6537)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6029..6048)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6708..7499,7500..7568))
                     /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     7331..7350
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7569..7673)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct 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 atgggtggag 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 gcttggtacc
      901 gagctcggat ccttgggtac ttccaatcca tttgatggcc gaggcccccc cgcgccgcct
      961 cggcctgggt cccccgcccg gggacgcccc ccgcgcggag ctagtggcgc tcacggccgt
     1021 gcagagcgaa cagggcgagg cgggcggggg cggctccccg cgccgcctcg gcctcctggg
     1081 cagccccctg ccgccgggcg cgcccctccc tgggccgggc tccggctcgg gctccgcctg
     1141 cggccagcgc tcctcggccg cgcacaagcg ctaccgccgc ctgcagaact gggtctacaa
     1201 cgtgctggag cggccccgcg gctgggcctt cgtctaccac gtcttcatat ttttgctggt
     1261 cttcagctgt ctggtgctgt ctgtgctgtc cactatccag gagcaccagg aacttgccaa
     1321 cgagtgtctc ctcatcttgg aattcgtgat gatcgtggtt ttcggcttgg agtacatcgt
     1381 ccgggtctgg tccgccggat gctgctgccg ctaccgagga tggcagggtc gcttccgctt
     1441 tgccagaaag cccttctgtg tcatcgactt catcgtgttc gtggcctcgg tggccgtcat
     1501 cgccgcgggt acccagggca acatcttcgc cacgtccgcg ctgcgcagca tgcgcttcct
     1561 gcagatcctg cgcatggtgc gcatggaccg ccgcggcggc acctggaagc tgctgggctc
     1621 agtggtctac gcgcatagca aggagctgat caccgcctgg tacatcgggt tcctggtgct
     1681 catcttcgcc tccttcctgg tctacctggc cgagaaggac gccaactccg acttctcctc
     1741 ctacgccgac tcgctctggt gggggacgat tacattgaca accatcggct atggtgacaa
     1801 gacaccgcac acatggctgg gcagggtcct ggctgctggc ttcgccttac tgggcatctc
     1861 tttctttgcc ctgcctgccg gcatcctagg ctccggcttt gccctgaagg tccaggagca
     1921 gcaccggcag aagcacttcg agaagcggag gatgccggca gccaacctca tccaggctgc
     1981 ctggcgcctg tactccaccg atatgagccg ggcctacctg acagccacct ggtacaagct
     2041 tgacgacatc atgcctgctg tgaagacagt catccgctcc atcaggattc tcaagttcct
     2101 ggtggccaaa aggaaattca aggagacact gcgaccgtac gacgtgaagg acgtcattga
     2161 gcagtactca gcaggccacc tggacatgct gggccggatc aagagcctgc aaactcgggt
     2221 ggaccaaatt gtgggtcggg ggcccgggga caggaaggcc cgggagaagg gcgacaaggg
     2281 gccctccgac gcggaggtgg tggatgaaat cagcatgatg ggacgcgtgg tcaaggtgga
     2341 gaagcaggtg cagtccatcg agcacaagct ggacctgctg ttgggcttct attcgcgcgc
     2401 cctgcgctaa ggcaaattgg gattaattaa tctcgaaaac ctgtacttcc aatccgtgag
     2461 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt
     2521 aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct
     2581 gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac
     2641 caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga
     2701 cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga
     2761 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg
     2821 catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga
     2881 gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa
     2941 ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta
     3001 ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag
     3061 cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga
     3121 gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagc atcaccatca
     3181 ccatcaccat cactaaggat cctgcagata tccatcacac tggcggccgc tcgagcatgc
     3241 atctagaggg ccctattcta tagtgtcacc taaatgctag agctcgctga tcagcctcga
     3301 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     3361 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     3421 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     3481 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     3541 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     3601 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     3661 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     3721 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     3781 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     3841 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     3901 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     3961 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     4021 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     4081 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     4141 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     4201 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     4261 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     4321 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     4381 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     4441 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     4501 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     4561 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
     4621 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     4681 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     4741 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     4801 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     4861 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     4921 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     4981 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     5041 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     5101 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     5161 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     5221 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     5281 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     5341 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
     5401 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     5461 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc
     5521 tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     5581 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     5641 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     5701 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     5761 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     5821 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     5881 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     5941 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     6001 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     6061 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     6121 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     6181 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     6241 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     6301 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     6361 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     6421 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     6481 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     6541 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     6601 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     6661 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     6721 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     6781 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     6841 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     6901 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     6961 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     7021 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     7081 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     7141 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     7201 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     7261 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     7321 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     7381 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     7441 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     7501 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     7561 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     7621 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     7681 ccccgaaaag tgccacctga cgtc
//
