LOCUS       sequence-349388-        6969 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses KCR1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6969)
  AUTHORS   Govorunova EG, Gou Y, Sineshchekov OA, Li H, Lu X, Wang Y, Brown LS,
            St-Pierre F, Xue M, Spudich JL
  TITLE     Kalium channelrhodopsins are natural light-gated potassium channels 
            that mediate optogenetic inhibition.
  JOURNAL   Nat Neurosci. 2022 Jun 20. pii: 10.1038/s41593-022-01094-6. doi: 
            10.1038/s41593-022-01094-6.
  PUBMED    35726059
REFERENCE   2  (bases 1 to 6969)
  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: "Nat 
            Neurosci. 2022 Jun 20. pii: 10.1038/s41593-022-01094-6. doi: 
            10.1038/s41593-022-01094-6."
COMMENT     Sequence Label: HcKCR1_mCherry_pcDNA3.1
FEATURES             Location/Qualifiers
     source          1..6969
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     58..77
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             212..1006
                     /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(266..285)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     876..895
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1182..1303
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1219..1238)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1273..1292
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1352..1368)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1352..1368)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1365..1387)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1376..1392
                     /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(1400..1406,1407..1424,1425..1430))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1445..1466
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1583..1600)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1754..2342)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1834..1853)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2513..3304,3305..3373))
                     /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     3136..3155
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3374..3478)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3553..3572)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3744..4123
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4124..4327
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4278..4298
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4372..4391
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4372..4390
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5239..5946
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(5381..5399)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5649..5668
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             5976..6017
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             6027..6044
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6067..6084)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6073..6297
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      6343..6771
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6430..6449)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6640..6661
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6780..6800)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        join(6785..6969,1..145)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6965..6969,1..131)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
       61 ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt
      121 ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc
      181 tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
      241 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
      301 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
      361 gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct
      421 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
      481 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
      541 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
      601 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
      661 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
      721 gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
      781 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
      841 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
      901 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
      961 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     1021 ttcgcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg
     1081 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     1141 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt
     1201 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     1261 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt
     1321 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     1381 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     1441 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     1501 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     1561 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     1621 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     1681 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     1741 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     1801 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     1861 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     1921 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     1981 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     2041 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     2101 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     2161 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     2221 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     2281 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     2341 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     2401 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     2461 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     2521 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     2581 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     2641 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     2701 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     2761 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     2821 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     2881 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     2941 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     3001 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     3061 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     3121 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     3181 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     3241 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     3301 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     3361 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     3421 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     3481 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
     3541 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
     3601 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg
     3661 cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga
     3721 tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat
     3781 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
     3841 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     3901 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
     3961 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
     4021 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
     4081 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
     4141 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
     4201 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
     4261 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
     4321 cagagctctc tggctaacta gagaacccac tgcttactgg cttatcgaaa ttaatacgac
     4381 tcactatagg gagacccaag ctggctagtt aagcttggta ccgagctcgg atccatgcct
     4441 ttctacgaca gccggccccc tgagggatgg cctaagggct ccatcaacga catggattat
     4501 ccactgctgg gctctatttg tgccgtgtgc tgcgtgttcg tggcaggcag cggcatctgg
     4561 atgctgtaca ggctggatct gggcatgggc tactcttgta agccatataa aagcggccgg
     4621 gcacctgagg tgaacagcct gtccggcatc atctgcctgc tgtgcggcac aatgtacgcc
     4681 gccaagtcct tcgacttctt tgatggcggc ggaaccccct tttctctgaa ttggtactgg
     4741 tatctggact acgtgttcac ctgccccctg ctgatcctgg acttcgcctt taccctggat
     4801 ctgcctcaca agatccggta cttctttgcc gtgtttctga ccctgtggtg tggagtggca
     4861 gccttcgtga caccaagcgc ctatagattt gcctactatg ccctgggctg ctgttggttc
     4921 acaccctttg ccctgtccct gatgcggcac gtgaaggaga gatacctggt gtatccaccc
     4981 aagtgccaga ggtggctgtt ctgggcctgc gtgatcttct ttggcttctg gcctatgttt
     5041 ccaatcctgt tcatcttttc ctggctgggc accggccaca tctctcagca ggccttctac
     5101 atcatccacg cctttctgga tctgacctgc aagtctatct tcggcatcct gatgacagtg
     5161 tttcgcctgg agctggagga gcacacagag gtgcagggac tgcccctgaa tgagcctgag
     5221 acactgtccg cggccgccgt gagcaagggc gaggaggata acatggccat catcaaggag
     5281 ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt cgagatcgag
     5341 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag
     5401 ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta cggctccaag
     5461 gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt ccccgagggc
     5521 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt gacccaggac
     5581 tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc
     5641 tccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctcctc cgagcggatg
     5701 taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct gaaggacggc
     5761 ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc
     5821 ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga ctacaccatc
     5881 gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac
     5941 aagtaacata tgctctagag ggcccgcggt tcgaaggtaa gcctatccct aaccctctcc
     6001 tcggtctcga ttctacgcgt accggtcatc atcaccatca ccattgagtt taaacccgct
     6061 gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc
     6121 cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg
     6181 catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca
     6241 agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt
     6301 ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg
     6361 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     6421 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     6481 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     6541 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     6601 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     6661 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     6721 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg
     6781 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     6841 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     6901 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     6961 gcccatccc
//
