LOCUS       40924_34754        5944 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pPKm-113, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5944)
  AUTHORS   Kyriakakis P, Catanho M, Hoffner N, Thavarajah W, Jian-Yu V, Chao 
            SS, Hsu A, Pham V, Naghavian L, Dozier LE, Patrick G, Coleman T.
  TITLE     Biosynthesis of Orthogonal Molecules Using Ferredoxin and 
            Ferredoxin-NADP+ Reductase Systems Enables Genetically Encoded PhyB 
            Optogenetics
  JOURNAL   ACS Synth Biol (2018) In press
  PUBMED    29301067
REFERENCE   2  (bases 1 to 5944)
  AUTHORS   Kyriakakis P.
  TITLE     Direct Submission
  JOURNAL   Submitted (15-NOV-2017) Bioengineering, UCSD, 9500 Gilman drive, 
            PFBH room 251 MC0412, La Jolla, CA 92093, United States of America
REFERENCE   3  (bases 1 to 5944)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5944)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "ACS Synth 
            Biol (2018) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (15-NOV-2017) Bioengineering, UCSD, 9500 Gilman drive, PFBH room 251
            MC0412, La Jolla, CA 92093, United States of America"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5944
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        27..406
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        407..610
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     promoter        655..673
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    705..880
                     /label=MTAD
                     /note="MTAD"
     CDS             885..896
                     /codon_start=1
                     /label=Factor Xa site
                     /note="Factor Xa recognition and cleavage site"
                     /translation="IEGR"
     CDS             912..932
                     /codon_start=1
                     /label=SV40 NLS
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /translation="PKKKRKV"
     misc_feature    977..1219
                     /label=PIF6
                     /note="PIF6"
     promoter        complement(1288..1306)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     polyA_signal    1332..1556
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1602..2030
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2044..2374
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             2441..3232
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    3409..3542
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3579..3595)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(3603..3619)
                     /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(3627..3657)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(3672..3693)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(3981..4569)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4743..5600)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5601..5705)
                     /label=AmpR promoter
ORIGIN
        1 cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc
       61 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
      121 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
      181 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg actatttacg
      241 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
      301 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
      361 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
      421 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
      481 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
      541 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
      601 aagcagagct ctctggctaa ctagagaacc cactgcttac tggcttatcg aaattaatac
      661 gactcactat agggagaccc aagcttggta ccgagctcgg atccatggga tctcctaggt
      721 ttgatactga cctgttcgat accgacttgt ttgatactga cctttttgat actgacctgt
      781 tcgataccga cttgtttgat actgaccttt ttgatactga cctgttcgat accgacttgt
      841 ttgatactga cctttttgat actgacctgt tcgataccga cttgatcgag ggaaggggtg
      901 gcgaattcat gcctaagaaa aagagaaagg tgggcggagg cgctagcatg atgttcttac
      961 caaccgatta ttgttgcagg ttaagcgatc aagagtatat ggagcttgtg tttgagaatg
     1021 gccagattct tgcaaagggc caaagatcca acgtttctct gcataatcaa cgtaccaaat
     1081 cgatcatgga tttgtatgag gcagagtata acgaggattt catgaagagt atcatccatg
     1141 gtggtggtgg tgccatcaca aatctcgggg acacgcaggt tgttccacaa agtcatgttg
     1201 ctgctgccca tgaaacaaac atgttggaaa gcaataaaca tgttgactga taagcggccg
     1261 ctcgagcatg catctagagg gccctattct atagtgtcac ctaaatgcta gagctcgctg
     1321 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     1381 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     1441 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     1501 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     1561 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     1621 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     1681 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     1741 tcaagctcta aatcggggca tccctttagg gttccgattt agtgctttac ggcacctcga
     1801 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     1861 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     1921 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tggggatttc
     1981 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     2041 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc caggcaggca gaagtatgca
     2101 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     2161 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     2221 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     2281 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
     2341 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
     2401 tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
     2461 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     2521 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     2581 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
     2641 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     2701 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     2761 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     2821 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     2881 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     2941 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
     3001 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     3061 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     3121 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     3181 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     3241 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     3301 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     3361 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
     3421 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     3481 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     3541 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
     3601 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     3661 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     3721 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     3781 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     3841 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     3901 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     3961 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     4021 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     4081 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     4141 gcctttctcc cttcgggaag cgtggcgctt tctcaatgct cacgctgtag gtatctcagt
     4201 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     4261 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     4321 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     4381 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     4441 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     4501 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     4561 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     4621 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     4681 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     4741 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     4801 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     4861 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     4921 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     4981 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     5041 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     5101 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     5161 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     5221 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     5281 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     5341 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     5401 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     5461 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     5521 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     5581 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     5641 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     5701 ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc
     5761 ccctatggtc gactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc
     5821 tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa
     5881 ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc
     5941 ttcg
//
