LOCUS       sequence-391080-        6388 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression vector for N-ter GFP11 and C-ter R9 tagged 
            WUSCHEL protein from Arabidopsis thaliana..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6388)
  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 6388)
  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: 1BR9-AtWUS
FEATURES             Location/Qualifiers
     source          1..6388
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     198..217
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     301..320
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        301..319
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    320..344
                     /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)."
     RBS             359..381
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             400..417
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             418..438
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             445..492
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     CDS             1552..1569
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1622..1640)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1636..1683
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1720..2175
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1807..1826)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2017..2038
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2268..3083)
                     /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             complement(2268..3083)
                     /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     2991..3010
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3075..3094
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3205..3793
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3694..3713
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3947..3964
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3979..4118
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4044..4063)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4204..4226
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4220..4411)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5220..6302)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     6264..6283
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6303..6380)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 cgacgctctc ccttatgcga ctcctgcatt aggaagcagc ccagtagtag gttgaggccg
       61 ttgagcaccg ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg
      121 gccacggggc ctgccaccat acccacgccg aaacaagcgc tcatgagccc gaagtggcga
      181 gcccgatctt ccccatcggt gatgtcggcg atataggcgc cagcaaccgc acctgtggcg
      241 ccggtgatgc cggccacgat gcgtccggcg tagaggatcg agatctcgat cccgcgaaat
      301 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
      361 tgtttaactt taagaaggag atataccatg ggttcttctc accatcacca tcaccatgaa
      421 aacctgtact tccaatccaa tgcacgcgat cacatggtcc tgcacgagta cgtgaacgcc
      481 gccgggatca ctggtggcgg aggttctgga ggcggtggat cgtacttcca atccaatgca
      541 gaacctcccc agcaccagca ccatcaccac caggctgatc aagagtcggg aaataacaac
      601 aacaataaaa gcgggagcgg gggttacacc tgtcgtcaaa catccacgcg ctggacacca
      661 accactgaac aaatcaaaat tttgaaagaa ttatactata acaatgccat tcgttcacct
      721 accgcagatc aaatccaaaa aattacagca cgcttgcgcc agttcggaaa gattgaggga
      781 aaaaacgtgt tctactggtt tcagaatcat aaggcacgtg aacgtcaaaa aaaacgtttt
      841 aacgggacta atatgacaac accgagttcg agtcccaact ccgttatgat ggcagcgaac
      901 gatcactatc atccattact gcaccaccat catggagtcc caatgcagcg tccagccaat
      961 tccgtgaatg tcaagctgaa ccaagaccac catctttatc accacaataa gccttaccca
     1021 tcttttaata acggcaacct gaatcacgcc agcagtggca ccgaatgcgg ggtcgtcaat
     1081 gcgtcgaatg gatatatgtc gtcgcacgtc tacggttcta tggaacaaga ctgttcaatg
     1141 aattacaaca acgtaggagg gggctgggcc aatatggacc atcactacag ctctgcgccg
     1201 tacaatttct tcgaccgtgc taaaccgtta ttcggcttag agggacatca ggaggaggag
     1261 gaatgcgggg gtgacgctta cttagaacat cgtcgtactt taccgctttt cccgatgcat
     1321 ggagaggacc atatcaatgg tgggtctggc gccatttgga agtacggaca gtcggaggta
     1381 cgcccttgcg caagtcttga gctgcgtttg aatggcattg gtagtgggag caacggcagc
     1441 agcggatccg tgagccgccg tcgccgtcgc cgtcgccgtc gctaataaca ttggaagtgg
     1501 ataacggatc cgaattcgag cgccgtcgac aagcttgcgg ccgcactcga gcaccaccac
     1561 caccaccact gagatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc
     1621 accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt
     1681 ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt agcggcgcat
     1741 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
     1801 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     1861 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     1921 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
     1981 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
     2041 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
     2101 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaactag
     2161 taacgtttac aatttcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     2221 tatttttcta aatacattca aatatgtatc cgctcatgaa ttaattctta gaaaaactca
     2281 tcgagcatca aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga
     2341 aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga
     2401 tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc
     2461 tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag
     2521 aatggcaaaa gtttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg
     2581 tcatcaaaat cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga
     2641 cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc
     2701 aggaacactg ccagcgcatc aacaatgttt tcacctgaat caggatattc ttctaatacc
     2761 tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg
     2821 ataaaatgct tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc
     2881 tcatctgtaa catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca
     2941 tcgggcttcc catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc
     3001 catttatacc catataaatc agcatccatg ttggaattta atcgcggcct agagcaagac
     3061 gtttcccgtt gaatatggct cataacaccc cttgtattac tgtttatgta agcagacagt
     3121 tttattgttc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     3181 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     3241 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     3301 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt
     3361 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     3421 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     3481 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     3541 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     3601 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     3661 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     3721 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     3781 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     3841 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     3901 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     3961 cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
     4021 acaccgcaat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
     4081 acactccgct atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg
     4141 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
     4201 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgaggcagc
     4261 tgcggtaaag ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg
     4321 cgtccagctc gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca
     4381 tgttaagggc ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt
     4441 tcatgggggt aatgataccg atgaaacgag agaggatgct cacgatacgg gttactgatg
     4501 atgaacatgc ccggttactg gaacgttgtg agggtaaaca actggcggta tggatgcggc
     4561 gggaccagag aaaaatcact cagggtcaat gccagcgctt cgttaataca gatgtaggtg
     4621 ttccacaggg tagccagcag catcctgcga tgcagatccg gaacataatg gtgcagggcg
     4681 ctgacttccg cgtttccaga ctttacgaaa cacggaaacc gaagaccatt catgttgttg
     4741 ctcaggtcgc agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt
     4801 cattctgcta accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca
     4861 cgatcatgcg cacccgtggg gccgccatgc cggcgataat ggcctgcttc tcgccgaaac
     4921 gtttggtggc gggaccagtg acgaaggctt gagcgagggc gtgcaagatt ccgaataccg
     4981 caagcgacag gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc
     5041 agagcgctgc cggcacctgt cctacgagtt gcatgataaa gaagacagtc ataagtgcgg
     5101 cgacgatagt catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg
     5161 gcatcggtcg agatcccggt gcctaatgag tgagctaact tacattaatt gcgttgcgct
     5221 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     5281 gcgcggggag aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag
     5341 acgggcaaca gctgattgcc cttcaccgcc tggccctgag agagttgcag caagcggtcc
     5401 acgctggttt gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa
     5461 catgagctgt cttcggtatc gtcgtatccc actaccgaga tatccgcacc aacgcgcagc
     5521 ccggactcgg taatggcgcg cattgcgccc agcgccatct gatcgttggc aaccagcatc
     5581 gcagtgggaa cgataccctc attcagcatt tgcatggttt gttgaaaacc ggacatggca
     5641 ctccagtcgc cttcccgttc cgctatcggc tgaatttgat tgcgagtgag atatttatgc
     5701 cagccagcca gacgcagacg cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt
     5761 tgctggtgac ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag
     5821 aaaataatac tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta
     5881 gtgcaggcag cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc
     5941 ccactgacgc gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt
     6001 cgttctacca tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc
     6061 gcgacaattt gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac
     6121 gactgtttgc ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc
     6181 gccgcttcca ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg
     6241 gaaacggtct gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc
     6301 acattcacca ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt
     6361 ttgcgccatt cgatggtgtc cgggatct
//
