LOCUS       40924_34864        6103 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pPKm-292, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6103)
  AUTHORS   Kyriakakis P, Catanho M, Hoffner N, Thavarajah W, Hu VJ, Chao SS, 
            Hsu A, Pham V, Naghavian L, Dozier LE, Patrick GN, Coleman TP.
  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 6103)
  AUTHORS   Kyriakakis P.
  TITLE     Direct Submission
  JOURNAL   Submitted (30-NOV-2017) Bioengineering, UCSD, 9500 Gilman Drive, 
            PFBH Room 251 MC0412, La Jolla, CA 92093, USA
REFERENCE   3  (bases 1 to 6103)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6103)
  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 
            (30-NOV-2017) Bioengineering, UCSD, 9500 Gilman Drive, PFBH Room 251
            MC0412, La Jolla, CA 92093, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6103
                     /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"
     CDS             696..716
                     /codon_start=1
                     /label=SV40 NLS
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /translation="PKKKRKV"
     CDS             726..1166
                     /codon_start=1
                     /label=GAL4 DNA binding domain
                     /note="DNA binding domain of the GAL4 transcriptional
                     activator"
                     /translation="MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTK
                     RSPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNK
                     DAVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVS"
     misc_feature    1182..1357
                     /label=MTAD
                     /note="MTAD"
     CDS             1362..1373
                     /codon_start=1
                     /label=Factor Xa site
                     /note="Factor Xa recognition and cleavage site"
                     /translation="IEGR"
     CDS             1389..1409
                     /codon_start=1
                     /label=SV40 NLS
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /translation="PKKKRKV"
     promoter        complement(1447..1465)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     polyA_signal    1491..1715
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1761..2189
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2203..2533
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             2600..3391
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    3568..3701
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3738..3754)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(3762..3778)
                     /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(3786..3816)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(3831..3852)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(4140..4728)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4902..5759)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5760..5864)
                     /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 ccatgcctaa gaaaaagaga aaggtgggcg
      721 gaggcatgaa gctactgtct tctatcgaac aagcatgcga tatttgccga cttaaaaagc
      781 tcaagtgctc caaagaaaaa ccgaagtgcg ccaagtgtct gaagaacaac tgggagtgtc
      841 gctactctcc caaaaccaaa aggtctccgc tgactagggc acatctgaca gaagtggaat
      901 caaggctaga aagactggaa cagctatttc tactgatttt tcctcgagaa gaccttgaca
      961 tgattttgaa aatggattct ttacaggata taaaagcatt gttaacagga ttatttgtac
     1021 aagataatgt gaataaagat gccgtcacag atagattggc ttcagtggag actgatatgc
     1081 ctctaacatt gagacagcat agaataagtg cgacatcatc atcggaagag agtagtaaca
     1141 aaggtcaaag acagttgact gtatcgccgg aattcggatc catgggatct cctaggtttg
     1201 atactgacct gttcgatacc gacttgtttg atactgacct ttttgatact gacctgttcg
     1261 ataccgactt gtttgatact gacctttttg atactgacct gttcgatacc gacttgtttg
     1321 atactgacct ttttgatact gacctgttcg ataccgactt gatcgaggga aggggtggcg
     1381 aattcatgcc taagaaaaag agaaaggtgg gcggaggcgc taggctagcc tgatagaggg
     1441 ccctattcta tagtgtcacc taaatgctag agctcgctga tcagcctcga ctgtgccttc
     1501 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     1561 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
     1621 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
     1681 tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg
     1741 gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     1801 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     1861 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcggggcat
     1921 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     1981 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     2041 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     2101 ggtctattct tttgatttat aagggatttt ggggatttcg gcctattggt taaaaaatga
     2161 gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt
     2221 ggaaagtccc caggctcccc aggcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2281 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     2341 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     2401 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     2461 cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     2521 aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcaagagac
     2581 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
     2641 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
     2701 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
     2761 cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg
     2821 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
     2881 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
     2941 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
     3001 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
     3061 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
     3121 caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
     3181 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
     3241 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     3301 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     3361 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc
     3421 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
     3481 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     3541 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa
     3601 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     3661 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag
     3721 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     3781 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     3841 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     3901 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     3961 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     4021 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     4081 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     4141 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     4201 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     4261 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     4321 gtggcgcttt ctcaatgctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     4381 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     4441 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     4501 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     4561 aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc
     4621 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     4681 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     4741 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     4801 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     4861 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     4921 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     4981 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     5041 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     5101 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     5161 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     5221 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     5281 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     5341 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     5401 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     5461 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     5521 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     5581 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     5641 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     5701 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     5761 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     5821 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     5881 gtgccacctg acgtcgacgg atcgggagat ctcccgatcc cctatggtcg actctcagta
     5941 caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg
     6001 tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaatt
     6061 gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct tcg
//
