LOCUS       sequence-197020-        3584 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Bacterial expression of the primed conversion capable protein 
            pr-mKikGR1 (mKikGR1-V69T)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3584)
  AUTHORS   Mohr MA, Kobitski AY, Sabater LR, Nienhaus K, Obara CJ, 
            Lippincott-Schwartz J, Nienhaus GU, Pantazis P
  TITLE     Rational Engineering of Photoconvertible Fluorescent Proteins for 
            Dual-Color Fluorescence Nanoscopy Enabled by a Triplet-State 
            Mechanism of Primed Conversion.
  JOURNAL   Angew Chem Int Ed Engl. 2017 Jun 29. doi: 10.1002/anie.201706121.
  PUBMED    28661566
REFERENCE   2  (bases 1 to 3584)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Angew Chem 
            Int Ed Engl. 2017 Jun 29. doi: 10.1002/anie.201706121."
COMMENT     Sequence Label: pRSET-pr-mKikGR1 (mKikGR1-V69T)
FEATURES             Location/Qualifiers
     source          1..3584
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             51..73
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             93..110
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             114..146
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(150..158,159..173)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             180..878
                     /codon_start=1
                     /product="monomeric photoconvertible Kikume green-to-red 
                     fluorescent protein"
                     /label=mKikGR1
                     /translation="MSVITSEMKIELRMEGSVNGHKFVITGKGSGRPYEGTQTVDLTVI
                     EGGPLPFAFDILTTAFHYGNRVFTEYPEEIVDYFKQSFPEGYSWERSMSYEDGGICLAT
                     NNITMKKDGSNTFVNEIRFDGTNFPANGPVMQRKTVKWEPSTEKMYVRDGVLKGDVEMA
                     LLLEGGGHYRCDFRTTYKAKKVVQLPDYHYVDHQMEITSHDKDYNKVKAYEHAKAYSGT
                     YRGAKYEFEA"
     primer_bind     complement(966..984)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      980..1027
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1124..1579
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1211..1230)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1421..1442
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1605..1709
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1710..1778,1779..2570)
                     /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     complement(1928..1947)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2741..3329
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3230..3249
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3483..3500
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 taatacgact cactataggg agaccacaac ggtttccctc tagaaataat tttgtttaac
       61 tttaagaagg agatatacat atgcggggtt ctcatcatca tcatcatcat ggtatggcta
      121 gcatgactgg tggacagcaa atgggtcggg atctgtacga cgatgacgat aaggatccca
      181 tgagtgtgat tacatcagaa atgaagatcg agctgcgtat ggaaggctct gtaaacgggc
      241 acaagttcgt gattacaggg aaaggaagtg gcagacctta cgagggaaca cagactgtgg
      301 acctgacagt catagagggc ggacctcttc cttttgcttt cgatatcctg acaacagcat
      361 tccattacgg caaccgggta tttaccgaat acccagaaga aatagtagac tacttcaagc
      421 agtcgtttcc tgagggttat tcttgggaac gaagcatgag ttacgaagac gggggaattt
      481 gcctcgccac aaacaatata acgatgaaga aagacggcag caacactttt gtcaatgaaa
      541 ttcgatttga tggtacaaac tttcctgcca atggtccagt tatgcagagg aagaccgtca
      601 aatgggagcc atccactgag aaaatgtatg tgcgtgatgg agtgctgaag ggtgatgttg
      661 aaatggctct gttgcttgaa ggaggtggcc attaccgatg tgacttcaga actacttaca
      721 aagcaaagaa ggttgtccag ttgccagact atcactacgt ggatcatcaa atggagataa
      781 caagccatga caaggattac aacaaggtta aggcgtatga acatgctaag gcttattccg
      841 ggacctacag gggcgccaag tatgaatttg aagcctaact cgagatctgc agctggtacc
      901 atggaattcg aagcttgatc cggctgctaa caaagcccga aaggaagctg agttggctgc
      961 tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg
     1021 ttttttgctg aaaggaggaa ctatatccgg atctggcgta atagcgaaga ggcccgcacc
     1081 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc
     1141 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     1201 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     1261 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     1321 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     1381 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     1441 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     1501 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     1561 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     1621 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     1681 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     1741 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     1801 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     1861 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     1921 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     1981 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     2041 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     2101 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     2161 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     2221 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     2281 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     2341 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     2401 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     2461 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     2521 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     2581 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     2641 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     2701 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     2761 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     2821 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     2881 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     2941 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     3001 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     3061 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     3121 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     3181 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     3241 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     3301 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     3361 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     3421 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     3481 gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg
     3541 cgcgttggcc gattcattaa tgcaggatct cgatcccgcg aaat
//
