LOCUS       40924_9866        6901 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Expression vector pcDNA-NES-PhoCl-mCherry, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6901)
  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) In press
  PUBMED    28288123
REFERENCE   2  (bases 1 to 6901)
  AUTHORS   Zhang W.
  TITLE     Direct Submission
  JOURNAL   Submitted (01-FEB-2017) Chemistry, University of Alberta, 11227 
            Saskatchewan Drive, Department of Chemistry, CCIS 4-140, Edmonton, 
            Alberta T6G2C5, Canada
REFERENCE   3  (bases 1 to 6901)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6901)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat. 
            Methods (2017) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (01-FEB-2017) Chemistry, University of Alberta, 11227 Saskatchewan 
            Drive, Department of Chemistry, CCIS 4-140, Edmonton, Alberta 
            T6G2C5, Canada"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Assembly Method       :: GeneStudio v. 2.2.0.0
            Sequencing Technology :: Illumina
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..6901
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             979..1704
                     /codon_start=1
                     /label=PhoCl
                     /note="photocleavable protein that separates into two
                     fragments upon exposure to violet light (Zhang et al., 
                     2017)"
                     /translation="VIPDYFKQSFPEGYSWERSMTYEDGGICIATNDITMEGDSFINKI
                     HFKGTNFPPNGPVMQKRTVGWEASTEKMYERDGVLKGDVKMKLLLKGGGHYRCDYRTTY
                     KVKQKPVKLPDYHFVDHRIEILSHDKDYNKVKLYEHAVARNSTDSMDELYKGGSGGMVS
                     KGEETITSVIKPDMKNKLRMEGNVNGHAFVIEGEGSGKPFEGIQTIDLEVKEGAPLPFA
                     YDILTTAFHYGNRVFTKYPR"
     CDS             1750..2454
                     /codon_start=1
                     /label=mBlueberry1
                     /note="mBlueberry1 is a basic (constitutively fluorescent)
                     blue fluorescent protein published in 2007, derived from 
                     Discosoma."
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     polyA_signal    2501..2725
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2771..3199
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        3213..3542
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             3609..4400
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    4577..4710
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4747..4763)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(4771..4787)
                     /label=lac operator
                     /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(4795..4825)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(4840..4861)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(5149..5734)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5908..6765)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6766..6870)
                     /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 gctggctagc
      901 gccaccatga acctggtgga cctgcagaag aagctggagg agctggagct ggacgagcag
      961 cagggaggcg ggggcagcgt gatccctgac tacttcaagc agagcttccc cgagggctac
     1021 agctgggagc gcagcatgac ctacgaggac ggcggcatct gcatcgccac caacgacatc
     1081 acaatggagg gggacagctt catcaacaag atccacttca agggcacgaa cttccccccc
     1141 aacggccccg tgatgcagaa gaggaccgtg ggctgggagg ccagcaccga gaagatgtac
     1201 gagcgcgacg gcgtgctgaa gggcgacgtg aagatgaagc tgctgctgaa gggcggcggc
     1261 cactatcgct gcgactaccg caccacctac aaggtcaagc agaagcccgt aaagctgccc
     1321 gactaccact tcgtggacca ccgcatcgag atcctgagcc acgacaagga ctacaacaag
     1381 gtgaagctgt acgagcacgc cgtggcccgc aactccaccg acagcatgga cgagctgtac
     1441 aagggtggca gcggtggcat ggtgagcaag ggcgaggaga ccattacaag cgtgatcaag
     1501 cctgacatga agaacaagct gcgcatggag ggcaacgtga acggccacgc cttcgtgatc
     1561 gagggcgagg gcagcggcaa gcccttcgag ggcatccaga cgattgattt ggaggtgaag
     1621 gagggcgccc cgctgccctt cgcctacgac atcctgacca ccgccttcca ctacggcaac
     1681 cgcgtgttca ccaagtaccc acggggaggt ggaggtaccg gaggcggggg cagcggaggc
     1741 gggggcagcg tgagcaaggg cgaggaggat aacatggcca tcatcaagga gttcatgcgc
     1801 ttcaaggtgc acatggaggg ctccgtgaac ggccacgagt tcgagatcga gggcgagggc
     1861 gagggccgcc cctacgaggg cacccagacc gccaagctga aggtgaccaa gggtggcccc
     1921 ctgcccttcg cctgggacat cctgtcccct cagttcatgt acggctccaa ggcctacgtg
     1981 aagcaccccg ccgacatccc cgactacttg aagctgtcct tccccgaggg cttcaagtgg
     2041 gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg tgacccagga ctcctccctg
     2101 caggacggcg agttcatcta caaggtgaag ctgcgcggca ccaacttccc ctccgacggc
     2161 cccgtaatgc agaagaagac catgggctgg gaggcctcct ccgagcggat gtaccccgag
     2221 gacggcgccc tgaagggcga gatcaagcag aggctgaagc tgaaggacgg cggccactac
     2281 gacgctgagg tcaagaccac ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac
     2341 aacgtcaaca tcaagttgga catcacctcc cacaacgagg actacaccat cgtggaacag
     2401 tacgaacgcg ccgagggccg ccactccacc ggcggcatgg acgagctgta caagtaactc
     2461 gagtctagag ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag
     2521 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
     2581 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
     2641 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat
     2701 gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg
     2761 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     2821 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     2881 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     2941 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     3001 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     3061 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     3121 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     3181 caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     3241 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     3301 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     3361 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
     3421 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
     3481 ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
     3541 aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca ggatgaggat
     3601 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
     3661 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
     3721 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
     3781 atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
     3841 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
     3901 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
     3961 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
     4021 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
     4081 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca
     4141 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
     4201 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     4261 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     4321 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     4381 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
     4441 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     4501 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     4561 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     4621 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     4681 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     4741 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     4801 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     4861 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     4921 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     4981 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     5041 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     5101 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     5161 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     5221 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     5281 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     5341 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     5401 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     5461 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     5521 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     5581 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     5641 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggttttt ttgtttgcaa
     5701 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     5761 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     5821 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     5881 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     5941 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     6001 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     6061 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     6121 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     6181 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     6241 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     6301 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     6361 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     6421 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     6481 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     6541 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     6601 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     6661 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     6721 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     6781 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     6841 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     6901 c
//
