LOCUS       sequence-372802-        4360 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient expression vector for SKLPT imaging for target 2 (Mpphot) 
            in plant cells (Negative control vector)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4360)
  AUTHORS   Konno R, Tanaka H, Kodama Y
  TITLE     SKLPT imaging: Efficient in vivo pre-evaluation of genome-editing 
            modules using fluorescent protein with peroxisome targeting signal.
  JOURNAL   Biochem Biophys Res Commun. 2018 Sep 3;503(1):235-241. doi: 
            10.1016/j.bbrc.2018.06.008. Epub 2018 Jun 12.
  PUBMED    29885839
REFERENCE   2  (bases 1 to 4360)
  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"; doi: 
            "10.1016/j.bbrc.2018.06.008"; journalName: "Biochem Biophys Res 
            Commun"; date: "2018-09-3- 3"; volume: "503"; issue: "1"; pages: 
            "235-241"
COMMENT     Sequence Label: pGWT35S-sfGFP-SKL-Target2 (Mpphot)
FEATURES             Location/Qualifiers
     source          1..4360
                     /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"
     promoter        96..440
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     383..403
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     misc_feature    460..515
                     /label=TMV Omega
                     /note="translational enhancer from the tobacco mosaic virus
                     5'-leader sequence (Gallie et al., 1988)"
     protein_bind    559..583
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(586..588,589..591,592..1302)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(631..652)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     1239..1260
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(1361..1385)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     terminator      1430..1680
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     promoter        complement(1745..1763)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1746..1763)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1781..1797)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1781..1797)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1794..1816)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1805..1821
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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(join(1829..1835,1836..1853,1854..1859))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1874..1895
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2012..2029)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2183..2771)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2263..2282)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2929..3744)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTSAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3652..3671
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3736..3755
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     3929..3947
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3985..4007)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4107..4126
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4320..4342
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4334..4351
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4335..4351
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 taatacgact cactataggg cgcggccgca gaattcgagc tcggtacccg ggatctcgag
       61 gccagatctc ccgggctcag aagaccgagg gctattgaga cttttcaaca aagggtaata
      121 tcgggaaacc tcctcggatt ccattgccca gctatctgtc acttatcgaa aggacagtag
      181 aaaaggaagg tggctcctac aaatgccatc attgcgataa aggaaaggct atcgttcaag
      241 atgcctctac cgacagtggt cccaaagatg gacccccacc cacgaggaac atcgtggaaa
      301 aagaagacgt tccaaccacg tcttcaaagc aagtggattg atgtgatatc tccactgacg
      361 taagggatga cgcacaatcc cactatcctt cgcaagaccc ttcctctata taaggaagtt
      421 catttcattt ggagaggaca ggcttcttga gatccttcaa caattaccaa caacaacaaa
      481 caacaaacaa cattacaatt actatttaca attacagaat tcgagctcgg taccaagctt
      541 ggaggctcag gaggctccac aagtttgtac aaaaaagcag gcttcatggt gagcaagggc
      601 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
      661 cacaagttca gcgtgagagg cgagggcgag ggcgatgcca ccaacggcaa gctgaccctg
      721 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
      781 acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc
      841 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tctctttcaa ggacgacggc
      901 acttacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
      961 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     1021 ttcaacagcc acaacgtcta tatcactgcc gacaagcaga agaacggcat caaggccaac
     1081 ttcaagatcc gccacaacgt tgaggacggc agcgtgcagc tcgccgacca ctaccagcag
     1141 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag
     1201 tccgttctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     1261 accgccgccg ggatcactct cggcatggac gagctgtaca aggaccggaa acacaccttc
     1321 gggcgccggg gctaggtcca aactctaagt ctaagctttg acccagcttt cttgtacaaa
     1381 gtggttggag gctcaggagg ctccccatgg gctagcacta gtatctagaa tcgttcaaac
     1441 atttggcaat aaagtttctt aagattgaat cctgttgccg gtcttgcgat gattatcata
     1501 taatttctgt tgaattacgt taagcatgta ataattaaca tgtaatgcat gacgttattt
     1561 atgagatggg tttttatgat tagagtcccg caattataca tttaatacgc gatagaaaac
     1621 aaaatatagc gcgcaaacta ggataaatta tcgcgcgcgg tgtcatctat gttactagat
     1681 gcggccgcaa ttgtggatcc gctctagagt cgacctgcag gcatgcaagc ttgcggccgc
     1741 gtattctata gtgtcaccta aatagcatgg cgtaatcatg gtcatagctg tttcctgtgt
     1801 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     1861 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     1921 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     1981 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     2041 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     2101 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     2161 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     2221 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     2281 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     2341 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     2401 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     2461 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     2521 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     2581 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     2641 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     2701 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     2761 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     2821 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     2881 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgttttaatt agaaaaactc
     2941 atcgagcatc aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg
     3001 aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag
     3061 atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc
     3121 ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga
     3181 gaatggcaat agcttatgca tttctttcca gacttgttca acaggccagc cattacgctc
     3241 gtcatcaaaa tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag
     3301 acgaaatacg cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg
     3361 caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac
     3421 ctggaatgct gttttccctg ggatcgcaga ggtgagtaac catgcatcat caggagtacg
     3481 gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat
     3541 ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc
     3601 atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc
     3661 ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc ttgagcaaga
     3721 cgtttcccgt tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacat
     3781 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     3841 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     3901 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc
     3961 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     4021 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     4081 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     4141 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa tattactaca ggcgccattc
     4201 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     4261 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
     4321 ccagtcacga cgttgtaaaa cgacggccag tgagctagtg
//
