LOCUS       sequence-176854-        6874 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express PhoCl-mCherry with C-term myc tag in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6874)
  AUTHORS   Zhang W, Lohman AW, Zhuravlova Y, Lu X, Wiens MD, Hoi H, Yaganoglu 
            S, Mohr MA, Kitova EN, Klassen JS, Pantazis P, Thompson RJ, Campbell
            RE
  TITLE     Optogenetic control with a photocleavable protein, PhoCl.
  JOURNAL   Nat Methods. 2017 Mar 13. doi: 10.1038/nmeth.4222.
  PUBMED    28288123
REFERENCE   2  (bases 1 to 6874)
  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 
            Methods. 2017 Mar 13. doi: 10.1038/nmeth.4222."
COMMENT     Sequence Label: pcDNA-PhoCl-mCherry-myc
FEATURES             Location/Qualifiers
     source          1..6874
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        116..495
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        496..699
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     650..670
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     744..763
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        744..762
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      782..791
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             800..1525
                     /codon_start=1
                     /product="photocleavable protein that separates into two 
                     fragments upon exposure to violet light (Zhang et al., 
                     2017)"
                     /label=PhoCl
                     /translation="VIPDYFKQSFPEGYSWERSMTYEDGGICIATNDITMEGDSFINKI
                     HFKGTNFPPNGPVMQKRTVGWEASTEKMYERDGVLKGDVKMKLLLKGGGHYRCDYRTTY
                     KVKQKPVKLPDYHFVDHRIEILSHDKDYNKVKLYEHAVARNSTDSMDELYKGGSGGMVS
                     KGEETITSVIKPDMKNKLRMEGNVNGHAFVIEGEGSGKPFEGIQTIDLEVKEGAPLPFA
                     YDILTTAFHYGNRVFTKYPR"
     CDS             1571..2275
                     /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(1713..1731)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1981..2000
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2276..2305
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(2346..2363)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2352..2576
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2622..3050
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2709..2728)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2919..2940
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3059..3079)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3064..3393
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3244..3379
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3306..3325
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3460..4254
                     /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(3514..3533)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4124..4143
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4428..4549
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4465..4484)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4519..4538
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4598..4614)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4598..4614)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4611..4633)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4622..4638
                     /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(4646..4652,4653..4670,4671..4676))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4691..4712
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4829..4846)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5000..5588)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5080..5099)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5759..6550,6551..6619))
                     /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     6382..6401
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6620..6724)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6799..6818)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
       61 cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgttgacat
      121 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
      181 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
      241 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
      301 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
      361 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
      421 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
      481 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
      541 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
      601 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
      661 ggtaggcgtg tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc
      721 actgcttact ggcttatcga aattaatacg actcactata gggagaccca agctggctag
      781 cgccaccatg ggaggcagcg tgatccctga ctacttcaag cagagcttcc ccgagggcta
      841 cagctgggag cgcagcatga cctacgagga cggcggcatc tgcatcgcca ccaacgacat
      901 cacaatggag ggggacagct tcatcaacaa gatccacttc aagggcacga acttcccccc
      961 caacggcccc gtgatgcaga agaggaccgt gggctgggag gccagcaccg agaagatgta
     1021 cgagcgcgac ggcgtgctga agggcgacgt gaagatgaag ctgctgctga agggcggcgg
     1081 ccactatcgc tgcgactacc gcaccaccta caaggtcaag cagaagcccg taaagctgcc
     1141 cgactaccac ttcgtggacc accgcatcga gatcctgagc cacgacaagg actacaacaa
     1201 ggtgaagctg tacgagcacg ccgtggcccg caactccacc gacagcatgg acgagctgta
     1261 caagggtggc agcggtggca tggtgagcaa gggcgaggag accattacaa gcgtgatcaa
     1321 gcctgacatg aagaacaagc tgcgcatgga gggcaacgtg aacggccacg ccttcgtgat
     1381 cgagggcgag ggcagcggca agcccttcga gggcatccag acgattgatt tggaggtgaa
     1441 ggagggcgcc ccgctgccct tcgcctacga catcctgacc accgccttcc actacggcaa
     1501 ccgcgtgttc accaagtacc cacggggagg tggaggtacc ggaggcgggg gcagcggagg
     1561 cgggggcagc gtgagcaagg gcgaggagga taacatggcc atcatcaagg agttcatgcg
     1621 cttcaaggtg cacatggagg gctccgtgaa cggccacgag ttcgagatcg agggcgaggg
     1681 cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca agggtggccc
     1741 cctgcccttc gcctgggaca tcctgtcccc tcagttcatg tacggctcca aggcctacgt
     1801 gaagcacccc gccgacatcc ccgactactt gaagctgtcc ttccccgagg gcttcaagtg
     1861 ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc gtgacccagg actcctccct
     1921 gcaggacggc gagttcatct acaaggtgaa gctgcgcggc accaacttcc cctccgacgg
     1981 ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc tccgagcgga tgtaccccga
     2041 ggacggcgcc ctgaagggcg agatcaagca gaggctgaag ctgaaggacg gcggccacta
     2101 cgacgctgag gtcaagacca cctacaaggc caagaagccc gtgcagctgc ccggcgccta
     2161 caacgtcaac atcaagttgg acatcacctc ccacaacgag gactacacca tcgtggaaca
     2221 gtacgaacgc gccgagggcc gccactccac cggcggcatg gacgagctgt acaaggaaca
     2281 aaaactcatc tcagaagagg atctgtaact cgagtctaga gggcccgttt aaacccgctg
     2341 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     2401 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     2461 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     2521 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     2581 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     2641 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     2701 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     2761 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     2821 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     2881 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     2941 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     3001 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     3061 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     3121 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     3181 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     3241 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3301 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     3361 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     3421 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     3481 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     3541 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     3601 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     3661 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     3721 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     3781 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     3841 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     3901 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     3961 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     4021 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     4081 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     4141 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     4201 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     4261 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     4321 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     4381 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     4441 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     4501 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     4561 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     4621 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     4681 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     4741 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     4801 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     4861 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     4921 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     4981 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     5041 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5101 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     5161 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     5221 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     5281 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     5341 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     5401 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     5461 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     5521 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     5581 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     5641 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     5701 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     5761 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     5821 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     5881 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     5941 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6001 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6061 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6121 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     6181 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     6241 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     6301 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     6361 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     6421 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     6481 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     6541 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     6601 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     6661 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     6721 cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc
     6781 cctatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct
     6841 ccctgcttgt gtgttggagg tcgctgagta gtgc
//
