LOCUS       sequence-395516-        6975 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses HcCCR_mCherry fusion in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6975)
  AUTHORS   Govorunova EG, Sineshchekov OA, Brown LS, Bondar AN, Spudich JL
  TITLE     Structural Foundations of Potassium Selectivity in 
            Channelrhodopsins.
  JOURNAL   mBio. 2022 Dec 20;13(6):e0303922. doi: 10.1128/mbio.03039-22. Epub 
            2022 Nov 22.
  PUBMED    36413022
REFERENCE   2  (bases 1 to 6975)
  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: "mBio."; 
            date: "2022-12-20"; volume: "13(6):e0303922. doi"; pages: " 
            10.1128/mbio.03039-22. Epub 2022 Nov 22"
COMMENT     Sequence Label: HcCCR_mCherry_pcDNA3.1
FEATURES             Location/Qualifiers
     source          1..6975
                     /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             1736..2443
                     /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(1878..1896)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2146..2165
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2473..2514
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2524..2541
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2564..2581)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2570..2794
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2840..3268
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2927..2946)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3137..3158
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3277..3297)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3282..3611
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3462..3597
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3524..3543
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3678..4472
                     /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(3732..3751)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4342..4361
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4648..4769
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4685..4704)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4739..4758
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4818..4834)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4818..4834)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4831..4853)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4842..4858
                     /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(4866..4872,4873..4890,4891..4896))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4911..4932
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5049..5066)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5220..5808)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5300..5319)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5979..6770,6771..6839))
                     /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     6602..6621
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6840..6944)
                     /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 gctggctagt
      901 taagcttggt accgagctcg gatccgccac catgcccttc tgcggcggcc ggcctgagga
      961 cggatggcac cacggcagca tccacgacat ggattaccca ctgctgggcg ctatggccgc
     1021 catctgtagc gtgtttatcg gaggctccgg agcatggatg ctgtacaggc tggatctggg
     1081 cctgggctat tcttgcaagc cacaccacag cggatacgca ccagaggcca acagcttctc
     1141 cgccctgtcc tgtctggtgt ctggcaccat ctatgccgcc aagacatttg acttctttga
     1201 tggcggcggc acccctttct cctttaattg gtactggtat ctggactacg tgttcacctg
     1261 ccctctgatc ctgctggatg tgctgtatac actggagatc ccacacaagc tgaggttcgt
     1321 gtttgccgtg atcatcaccc tgtggtgcgg agtggcagcc ttcgtgacac catctgcctt
     1381 ccgctttggc tactatgccg tgggctgcgt gtggtttgtg cctttctctt ttagcctgct
     1441 gcggcacgtg aagcagagat accaggtgta tccccctaag tgccagaagc tgctgttctg
     1501 ggcctgtacc atcttctttg gcttctggcc actgtttccc atcctgttcc tgttttcctg
     1561 gctgggcacc ggacacatcg accagcaggc cttcacaatc atccacgcct tcctggatct
     1621 gttttgtaag accgtgtttg gcctgatcat gacattcttt cggctggagc tggaggagca
     1681 cacagaggtg ctgggcctgc ctctgaacga gccaaagggc aagcacgcgg ccgccgtgag
     1741 caagggcgag gaggataaca tggccatcat caaggagttc atgcgcttca aggtgcacat
     1801 ggagggctcc gtgaacggcc acgagttcga gatcgagggc gagggcgagg gccgccccta
     1861 cgagggcacc cagaccgcca agctgaaggt gaccaagggt ggccccctgc ccttcgcctg
     1921 ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc tacgtgaagc accccgccga
     1981 catccccgac tacttgaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa
     2041 cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg acggcgagtt
     2101 catctacaag gtgaagctgc gcggcaccaa cttcccctcc gacggccccg taatgcagaa
     2161 gaagaccatg ggctgggagg cctcctccga gcggatgtac cccgaggacg gcgccctgaa
     2221 gggcgagatc aagcagaggc tgaagctgaa ggacggcggc cactacgacg ctgaggtcaa
     2281 gaccacctac aaggccaaga agcccgtgca gctgcccggc gcctacaacg tcaacatcaa
     2341 gttggacatc acctcccaca acgaggacta caccatcgtg gaacagtacg aacgcgccga
     2401 gggccgccac tccaccggcg gcatggacga gctgtacaag taacatatgc tctagagggc
     2461 ccgcggttcg aaggtaagcc tatccctaac cctctcctcg gtctcgattc tacgcgtacc
     2521 ggtcatcatc accatcacca ttgagtttaa acccgctgat cagcctcgac tgtgccttct
     2581 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     2641 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     2701 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     2761 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     2821 ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     2881 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     2941 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     3001 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     3061 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     3121 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     3181 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     3241 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     3301 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     3361 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3421 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     3481 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     3541 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     3601 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat caagagacag
     3661 gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt
     3721 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     3781 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     3841 gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     3901 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     3961 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     4021 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     4081 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     4141 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     4201 aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga
     4261 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     4321 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     4381 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     4441 ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg cgaaatgacc
     4501 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
     4561 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     4621 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa
     4681 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     4741 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag
     4801 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     4861 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     4921 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     4981 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     5041 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     5101 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     5161 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     5221 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     5281 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     5341 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     5401 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     5461 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     5521 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     5581 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     5641 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     5701 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     5761 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     5821 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     5881 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     5941 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     6001 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     6061 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     6121 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     6181 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     6241 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     6301 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     6361 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     6421 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     6481 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     6541 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     6601 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     6661 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     6721 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     6781 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     6841 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     6901 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     6961 gtgccacctg acgtc
//
