LOCUS       sequence-197025-        3541 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of the primed conversion capable protein 
            pr-Kaede (Kaede-A69T)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3541)
  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 3541)
  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: "Angew Chem 
            Int Ed Engl. 2017 Jun 29. doi: 10.1002/anie.201706121."
COMMENT     Sequence Label: pRSET-pr-Kaede (Kaede-A69T)
FEATURES             Location/Qualifiers
     source          1..3541
                     /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..860
                     /codon_start=1
                     /product="Kaede green-to-red photoconvertible fluorescent 
                     protein (Ando et al., 2002)"
                     /label=Kaede
                     /translation="MVSLIKPEMKIKLLMEGNVNGHQFVIEGDGKGHPFEGKQSMDLVV
                     KEGAPLPFAYDILTTAFHYGNRVFTKYPDHIPDYFKQSFPKGFSWERSLMFEDGGVCIA
                     TNDITLKGDTFFNKVRFDGVNFPPNGPVMQKKTLKWEASTEKMYLRDGVLTGDITMALL
                     LKGDVHYRCDFRTTYKSRQEGVKLPGYHFVDHCISILRHDKDYNEVKLYEHAVAHSGLP
                     DNVK"
     primer_bind     complement(923..941)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      937..984
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1081..1536
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1168..1187)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1378..1399
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1562..1666
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1667..1735,1736..2527)
                     /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(1885..1904)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2698..3286
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3187..3206
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3440..3457
                     /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 aaggatccga
      181 tggtgagtct gattaaacca gaaatgaaga tcaagctgct tatggaaggc aatgtaaacg
      241 ggcaccagtt tgttattgag ggagatggaa aaggccatcc ttttgaggga aaacagagta
      301 tggaccttgt agtcaaagaa ggcgcacctc tcccttttgc ctacgatatc ttgacaacag
      361 cattccatta tggtaacagg gtttttacca aatacccaga ccatatacca gactacttca
      421 agcagtcgtt tcccaaaggg ttttcttggg agcgaagcct gatgttcgag gacgggggcg
      481 tttgcatcgc tacaaatgac ataacactga aaggagacac tttttttaac aaagttcgat
      541 ttgatggcgt aaactttccc ccaaatggtc ctgttatgca gaagaagact ctgaaatggg
      601 aggcatccac tgagaaaatg tatttgcgtg atggagtgtt gacgggcgat attaccatgg
      661 ctctgctgct taaaggagat gtccattacc gatgtgactt cagaactact tacaaatcta
      721 ggcaggaggg tgtcaagttg ccaggatatc actttgtcga tcactgcatc agcatattga
      781 ggcatgacaa agactacaac gaggttaagc tgtatgagca tgctgttgcc cattctggat
      841 tgccggacaa cgtcaagtaa gaattcgaag cttgatccgg ctgctaacaa agcccgaaag
      901 gaagctgagt tggctgctgc caccgctgag caataactag cataacccct tggggcctct
      961 aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggatc tggcgtaata
     1021 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggg
     1081 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     1141 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     1201 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     1261 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     1321 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     1381 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     1441 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     1501 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
     1561 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     1621 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     1681 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     1741 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     1801 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     1861 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     1921 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     1981 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     2041 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     2101 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     2161 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     2221 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     2281 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     2341 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     2401 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     2461 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     2521 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     2581 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     2641 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     2701 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     2761 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     2821 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     2881 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     2941 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     3001 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     3061 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     3121 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     3181 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     3241 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     3301 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     3361 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     3421 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     3481 caaaccgcct ctccccgcgc gttggccgat tcattaatgc aggatctcga tcccgcgaaa
     3541 t
//
