LOCUS       sequence-391139-        4957 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes SV40-mCherry-GFP1-10 in pDGB1-aR backbone for plant 
            expression. Requires pSOUP for replication in agrobacterium..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4957)
  AUTHORS   Wang JW, Squire HJ, Goh NS, Ni HM, Lien E, Wong C, Gonzalez-Grandio 
            E, Landry MP
  TITLE     Delivered complementation in planta (DCIP) enables measurement of 
            peptide-mediated protein delivery efficiency in plants.
  JOURNAL   Commun Biol. 2023 Aug 12;6(1):840. doi: 10.1038/s42003-023-05191-5.
  PUBMED    37573467
REFERENCE   2  (bases 1 to 4957)
  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: "Commun 
            Biol."; date: "2023-08-12"; volume: "6(1):840. doi"; pages: " 
            10.1038/s42003-023-05191-5"
COMMENT     Sequence Label: DCIP
FEATURES             Location/Qualifiers
     source          1..4957
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      672..1260
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1161..1180
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    1351..1375
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     promoter        1396..1741
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     1684..1704
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     misc_feature    1761..1816
                     /label=TMV Omega
                     /note="translational enhancer from the tobacco mosaic virus
                     5'-leader sequence (Gallie et al., 1988)"
     CDS             1838..1858
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1862..2566
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(2004..2022)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2272..2291
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2636..2664)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     terminator      3274..3526
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     primer_bind     complement(3561..3579)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    3768..3790
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA 
                     (truncated)"
     rep_origin      3917..4352
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
     primer_bind     complement(4632..4651)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             join(4643..4957,1..501)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             join(4643..4957,1..501)
                     /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
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4716..4735)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 gcattccagg tattagaaga atatcctgat tcaggtgaaa atattgttga tgcgctggca
       61 gtgttcctgc gccggttgca ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc
      121 gtatttcgac tcgctcaggc gcaatcacga atgaataacg gtttggttga tgcgagtgat
      181 tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat gcataaactt
      241 ttgccattct caccggattc agtcgtcact catggtgatt tctcacttga taaccttatt
      301 tttgacgagg ggaaattaat aggttgtatt gatgttggac gagtcggaat cgcagaccga
      361 taccaggatc ttgccatcct atggaactgc ctcggtgagt tttctccttc attacagaaa
      421 cggctttttc aaaaatatgg tattgataat cctgatatga ataaattgca gtttcatttg
      481 atgctcgatg agtttttcta atcactagac caatgttaca catatatact ttagattgat
      541 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
      601 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
      661 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
      721 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct tcttccgaag
      781 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
      841 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
      901 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
      961 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     1021 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     1081 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     1141 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     1201 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     1261 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     1321 agatctcaaa caaacacata cagcgactta gtttacccgc caatatatcc tgtcaggatc
     1381 cgtctcatga cggagtgaga cttttcaaca aagggtaata tcgggaaacc tcctcggatt
     1441 ccattgccca gctatctgtc acttcatcga aaggacagta gaaaaggaag gtggctccta
     1501 caaatgccat cattgcgata aaggaaaggc tatcgttcaa gatgcctcta ccgacagtgg
     1561 tcccaaagat ggacccccac ccacgaggaa catcgtggaa aaagaagatg ttccaaccac
     1621 gtcctcaaag caagtggatt gatgtgatat ctccactgac gtaagggatg acgcacaatc
     1681 ccactatcct tcgcaagacc cttcctctat ataaggaagt tcatttcatt tggagaggac
     1741 aggcttcttg agatccttca acaattacca acaacaacaa acaacaaaca acattacaat
     1801 tactatttac aattacagtc gactctagag gataatgcca aagaaaaagc ggaaagtcgg
     1861 agtgagcaag ggcgaggagg ataacatggc catcatcaag gagttcatgc gcttcaaggt
     1921 gcacatggag ggctccgtga acggccacga gttcgagatc gagggcgagg gcgagggccg
     1981 cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc ccctgccctt
     2041 cgcctgggac atcctgtccc ctcagttcat gtacggctcc aaggcctacg tgaagcaccc
     2101 cgccgacatc cccgactact tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt
     2161 gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc tccaggacgg
     2221 cgagttcatc tacaaggtga agctgcgcgg caccaacttc ccctccgacg gccccgtaat
     2281 gcagaagaag accatgggct gggaggcctc ctccgagcgg atgtaccccg aggacggcgc
     2341 cctgaagggc gagatcaagc agaggctgaa gctgaaggac ggcggccact acgacgctga
     2401 ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct acaacgtcaa
     2461 catcaagttg gacatcacct cccacaacga ggactacacc atcgtggaac agtacgaacg
     2521 cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaagggcg gtggcggtag
     2581 cggcggtggc agcctgcaga tgatcgatag caaaggagaa gaacttttca ctggagttgt
     2641 cccaattctt gttgaattag atggtgatgt taatgggcac aaattttctg tccgtggaga
     2701 gggtgaaggt gatgctacaa ttggaaaact cacccttaaa tttatttgca ctactggaaa
     2761 actacctgtt ccatggccaa cacttgtcac tactctgacc tatggtgttc aatgcttttc
     2821 ccgttatccg gatcacatga aacggcatga ctttttcaag agtgccatgc ccgaaggtta
     2881 tgtacaggaa cgcactatat ctttcaaaga tgacgggaaa tacaagacgc gtgctgtcgt
     2941 caagtttgaa ggtgataccc ttgttaatcg tatcgagtta aaaggtactg attttaaaga
     3001 agatggaaac attctcggac acaaactcga gtacaacttt aactcacaca atgtatacat
     3061 cacggcagac aaacaaaaga atggaatcaa agctaacttc accgttcgcc acaacgttga
     3121 agatggttcc gttcaactag cagaccatta tcaacaaaat actccaattg gcgatggccc
     3181 tgtcctttta ccagacaacc attacctgtc gacacaaact gtcctttcga aagatcccaa
     3241 cgaaaagtga gcttggaatg gatcttcgat cccgatcgtt caaacatttg gcaataaagt
     3301 ttcttaagat tgaatcctgt tgccggtctt gcgacgatta tcatataatt tctgttgaat
     3361 tacgttaagc atgtaataat taacatgtaa tgcatgacgt tatttatgag atgggttttt
     3421 atgattagag tcccgcaatt atacatttaa tacgcgatag aaaacaaaat atagcgcgca
     3481 aactaggata aattatcgcg cgcggtgtca tctatgttac tagatcggga attgccaagc
     3541 taattcttga agatgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     3601 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     3661 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     3721 aatgcttcaa taatgggacc gactcgcgct ctcctgagac ggatccgtta caccacaata
     3781 tatcctgcca agatctaatt ccggggatcg gaaatccaga agcccgagag gttgccgcct
     3841 ttcgggcttt ttctttttca aaaaaaaaaa tttataaaac gatctgttgc ggccggccgc
     3901 cgggttgtgg gcaaaggcgc tcgacggtgg gcaaccgctt gcggttgtcc acgggcggag
     3961 ccggtgcgcg tagcgcattg tccacaagcc aagggcgacc aataattgat atatatattc
     4021 ataattgaaa agctaattga acatactact tgctgtaact acttgccgga gcgaggggtg
     4081 tttgcaagct gttgatctga aagggctatt agcgttctca cgtgcctttt tgattagcga
     4141 tttcacgtga ccttattagc gatttcacgt actccgatta gcgatttcac gtaccctgat
     4201 tagcgatttc acgtggatag tttttggagc gggccggaaa gccccgtgaa tcaaggcttt
     4261 gcggggcatt agcggtttca cgtggataac taccctctat ccacaggctt ccggggataa
     4321 aaaagcccgc tcgacggcgg gctgttggat ggggatcgcc tgaatcgccc catcatccag
     4381 ccagaaagtg agggagccac ggttgatgag agctttgttg taggtggacc agttggtgat
     4441 tttgaacttt tgctttgcca cggaacggtc tgcgttgtcg ggaagatgcg tgatctgatc
     4501 cttcaactca gcaaaagttc gatttattca acaaagccac gttgtgtctc aaaatctctg
     4561 atgttacatt gcacaagata aaaatatatc atcatgaaca ataaaactgt ctgcttacat
     4621 aaacagtaat acaaggggtg ttatgagcca tattcaacgg gaaacgtctt gctcaaggcc
     4681 gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc gcgataatgt
     4741 cgggcaatca ggtgcgacaa tctaccgatt gtatgggaag cccgatgcgc cagagttgtt
     4801 tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg tcagactaaa
     4861 ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta ctcctgatga
     4921 tgcatggtta ctcaccactg cgatcccagg gaaaaca
//
