LOCUS       sequence-401532-        7474 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used to produce and express in planta N-terminal fusion proteins 
            with the GAL4 BD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7474)
  AUTHORS   Chen J, Luo M, Hands P, Rolland V, Zhang J, Li Z, Outram M, Dodds P,
            Ayliffe M
  TITLE     A split GAL4 RUBY assay for visual in planta detection of 
            protein-protein interactions.
  JOURNAL   Plant J. 2023 Jun;114(6):1209-1226. doi: 10.1111/tpj.16234. Epub 
            2023 Apr 26.
  PUBMED    37323061
REFERENCE   2  (bases 1 to 7474)
  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: "10.1111/tpj.16234";
            journalName: "Plant J"; date: "2023-06"; volume: "114"; issue: "6"; 
            pages: "1209-1226"
COMMENT     Sequence Label: 35S-GWY-GAL4BD/pAM
FEATURES             Location/Qualifiers
     source          1..7474
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(101..353)
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     CDS             complement(373..924)
                     /codon_start=1
                     /gene="Streptomyces hygroscopicus bar"
                     /product="phosphinothricin acetyltransferase"
                     /label=BlpR
                     /note="confers resistance to bialophos or phosphinothricin"
                     /translation="MSPERRPADIRRATEADMPAVCTIVNHYIETSTVNFRTEPQEPQE
                     WTDDLVRLRERYPWLVAEVDGEVAGIAYAGPWKARNAYDWTAESTVYVSPRHQRTGLGS
                     TLYTHLLKSLEAQGFKSVVAVIGLPNDPSVRMHEALGYAPRGMLRAAGFKHGNWHDVGF
                     WQLDFSLPVPPRPVLPVTEI"
     promoter        complement(966..1145)
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     promoter        1614..1958
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     1902..1922
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     primer_bind     1960..1976
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1961..1977
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1988..2112
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2137..2142,2143..2160,2161..2167)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2221..2880
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2287..2306)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3222..3527
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3406..3425
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(3568..3692)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             3739..4179
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="DNA binding domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 DNA binding domain
                     /translation="MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTK
                     RSPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNK
                     DAVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVS"
     misc_feature    4516..4540
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     rep_origin      4652..5361
                     /label=oriV
                     /note="incP origin of replication"
     promoter        5434..5525
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5526..5594,5595..6386)
                     /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(5744..5763)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6557..7145
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7046..7065
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    join(7457..7474,1..7)
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
ORIGIN
        1 tgtaaacaaa ttgacgctta gacaacttaa taacacattg cggacgtttt taatgatcga
       61 atactaacgt ctctaccaga tatcagcttg catgccggtc gatctagtaa catagatgac
      121 accgcgcgcg ataatttatc ctagtttgcg cgctatattt tgttttctat cgcgtattaa
      181 atgtataatt gcgggactct aatcataaaa acccatctca taaataacgt catgcattac
      241 atgttaatta ttacatgctt aacgtaattc aacagaaatt atatgataat catcgcaaga
      301 ccggcaacag gattcaatct taagaaactt tattgccaaa tgtttgaacg atctgcttga
      361 ctctaggggt catcagattt cggtgacggg caggaccgga cggggcggca ccggcaggct
      421 gaagtccagc tgccagaaac ccacgtcatg ccagttcccg tgcttgaagc cggccgcccg
      481 cagcatgccg cggggggcat atccgagcgc ctcgtgcatg cgcacgctcg ggtcgttggg
      541 cagcccgatg acagcgacca cgctcttgaa gccctgtgcc tccagggact tcagcaggtg
      601 ggtgtagagc gtggagccca gtcccgtccg ctggtggcgg ggggagacgt acacggtcga
      661 ctcggccgtc cagtcgtagg cgttgcgtgc cttccaggga cccgcgtagg cgatgccggc
      721 gacctcgccg tccacctcgg cgacgagcca gggatagcgc tcccgcagac ggacgaggtc
      781 gtccgtccac tcctgcggtt cctgcggctc ggtacggaag ttgaccgtgc ttgtctcgat
      841 gtagtggttg acgatggtgc agaccgccgg catgtccgcc tcggtggcac ggcggatgtc
      901 ggccgggcgt cgttctgggc tcatggtaga tcccctcgat cgagttgaga gtgaatatga
      961 gactctaatt ggataccgag gggaatttat ggaacgtcag tggagcattt ttgacaagaa
     1021 atatttgcta gctgatagtg accttaggcg acttttgaac gcgcaataat ggtttctgac
     1081 gtatgtgctt agctcattaa actccagaaa cccgcggctc agtggctcct tcaacgttgc
     1141 ggttctgtca gttccaaacg taaaacggct tgtcccgcgt catcggcggg ggtcataacg
     1201 tgactccctt aattctccgc tcatgatcag attgtcgttt cccgccttcg gtttgggcgc
     1261 gcccggttca gaagaccaga gggctattga gacttttcaa caaagggtaa tatcgggaaa
     1321 cctcctcgga ttccattgcc cagctatctg tcacttcatc gaaaggacag tagaaaagga
     1381 agatggcttc tacaaatgcc atcattgcga taaaggaaag gctatcgttc aagatgcctc
     1441 taccgacagt ggtcccaaag atggaccccc acccacgagg aacatcgtgg aaaaagaaga
     1501 cgttccaacc acgtcttcaa agcaagtgga ttgatgtgat acatggtgga gcacgacact
     1561 ctcgtctact ccaagaatat caaagataca gtctcagaag accagagggc tattgagact
     1621 tttcaacaaa gggtaatatc gggaaacctc ctcggattcc attgcccagc tatctgtcac
     1681 ttcatcgaaa ggacagtaga aaaggaagat ggcttctaca aatgccatca ttgcgataaa
     1741 ggaaaggcta tcgttcaaga tgcctctacc gacagtggtc ccaaagatgg acccccaccc
     1801 acgaggaaca tcgtggaaaa agaagacgtt ccaaccacgt cttcaaagca agtggattga
     1861 tgtgatatct ccactgacgt aagggatgac gcacaatccc actatccttc gcaagaccct
     1921 tcctctatat aaggaagttc atttcatttg gagaggacct cgaggtcgac ggtatcgata
     1981 agctatcaca agtttgtaca aaaaagctga acgagaaacg taaaatgata taaatatcaa
     2041 tatattaaat tagattttgc ataaaaaaca gactacataa tactgtaaaa cacaacatat
     2101 ccagtcacta tggcggccgc attaggcacc ccaggcttta cactttatgc ttccggctcg
     2161 tataatgtgt ggattttgag ttaggatccg tcgagatttt caggagctaa ggaagctaaa
     2221 atggagaaaa aaatcactgg atataccacc gttgatatat cccaatggca tcgtaaagaa
     2281 cattttgagg catttcagtc agttgctcaa tgtacctata accagaccgt tcagctggat
     2341 attacggcct ttttaaagac cgtaaagaaa aataagcaca agttttatcc ggcctttatt
     2401 cacattcttg cccgcctgat gaatgctcat ccggaattcc gtatggcaat gaaagacggt
     2461 gagctggtga tatgggatag tgttcaccct tgttacaccg ttttccatga gcaaactgaa
     2521 acgttttcat cgctctggag tgaataccac gacgatttcc ggcagtttct acacatatat
     2581 tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt tccctaaagg gtttattgag
     2641 aatatgtttt tcgtctcagc caatccctgg gtgagtttca ccagttttga tttaaacgtg
     2701 gccaatatgg acaacttctt cgcccccgtt ttcaccatgg gcaaatatta tacgcaaggc
     2761 gacaaggtgc tgatgccgct ggcgattcag gttcatcatg ccgtttgtga tggcttccat
     2821 gtcggcagaa tgcttaatga attacaacag tactgcgatg agtggcaggg cggggcgtaa
     2881 acgcgtggat ccggcttact aaaagccaga taacagtatg cgtatttgcg cgctgatttt
     2941 tgcggtataa gaatatatac tgatatgtat acccgaagta tgtcaaaaag aggtatgcta
     3001 tgaagcagcg tattacagtg acagttgaca gcgacagcta tcagttgctc aaggcatata
     3061 tgatgtcaat atctccggtc tggtaagcac aaccatgcag aatgaagccc gtcgtctgcg
     3121 tgccgaacgc tggaaagcgg aaaatcagga agggatggct gaggtcgccc ggtttattga
     3181 aatgaacggc tcttttgctg acgagaacag gggctggtga aatgcagttt aaggtttaca
     3241 cctataaaag agagagccgt tatcgtctgt ttgtggatgt acagagtgat attattgaca
     3301 cgcccgggcg acggatggtg atccccctgg ccagtgcacg tctgctgtca gataaagtct
     3361 cccgtgaact ttacccggtg gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg
     3421 atatggccag tgtgccggtc tccgttatcg gggaagaagt ggctgatctc agccaccgcg
     3481 aaaatgacat caaaaacgcc attaacctga tgttctgggg aatataaatg tcaggctccc
     3541 ttatacacag ccagtctgca ggtcgaccat agtgactgga tatgttgtgt tttacagtat
     3601 tatgtagtct gttttttatg caaaatctaa tttaatatat tgatatttat atcattttac
     3661 gtttctcgtt cagctttctt gtacaaagtg gtgatcggtg gtggtagtgg tggtggtgga
     3721 tcaggtggtg gtggatctat gaagctactg tcttctatcg aacaagcatg cgatatttgc
     3781 cgacttaaaa agctcaagtg ctccaaagaa aaaccgaagt gcgccaagtg tctgaagaac
     3841 aactgggagt gtcgctactc tcccaaaacc aaaaggtctc cgctgactag ggcacatctg
     3901 acagaagtgg aatcaaggct agaaagactg gaacagctat ttctactgat ttttcctcga
     3961 gaagaccttg acatgatttt gaaaatggat tctttacagg atataaaagc attgttaaca
     4021 ggattatttg tacaagataa tgtgaataaa gatgccgtca cagatagatt ggcttcagtg
     4081 gagactgata tgcctctaac attgagacag catagaataa gtgcgacatc atcatcggaa
     4141 gagagtagta acaaaggtca aagacagttg actgtatcgt aagaattcaa ggcctctcga
     4201 gcctctagaa ctatagtgag tcgtattact agagtccgca aaaatcacca gtctctctct
     4261 acaaatctat ctctctctat ttttctccag aataatgtgt gagtagttcc cagataaggg
     4321 aattagggtt cttatagggt ttcgctcatg tgttgagcat ataagaaacc cttagtatgt
     4381 atttgtattt gtaaaatact tctatcaata aaatttctaa ttcctaaaac caaaatccag
     4441 tgaccgggcg gccgccaccg cggtggaggg ggatcagatt gtcgtttccc gccttcagtt
     4501 taaactatca gtgtttgaca ggatatattg gcgggtaaac ctaagagaaa agagcgttta
     4561 ttagaataat cggatattta aaagggcgtg aaaaggttta tccgttcgtc catttgtatg
     4621 tgtacatcac cgacgagcaa ggcaagaccg agcgcctttc cgacgctcac cgggctggtt
     4681 gccctcgccg ctgggctggc ggccgtctat ggccctgcaa acgcgccaga aacgccgtcg
     4741 aagccgtgtg cgagacaccg gccgccggcg ttgtggatac ctcgcggaaa acttggccct
     4801 cactgacaga tgaggggcgg acgttgacac ttgaggggcc gactcacccg gcgcggcgtt
     4861 gacagatgag gggcaggctc gatttcggcc ggcgacgtgg agctggccag cctcgcaaat
     4921 cggcgaaaac gcctgatttt acgcgagttt cccacagatg atgtggacaa gcctggggat
     4981 aagtgccctg cggtattgac acttgagggg cgcgactact gacagatgag gggcgcgatc
     5041 cttgacactt gaggggcaga gtgctgacag atgaggggcg cacctattga catttgaggg
     5101 gctgtccaca ggcagaaaat ccagcatttg caagggtttc cgcccgtttt tcggccaccg
     5161 ctaacctgtc ttttaacctg cttttaaacc aatatttata aaccttgttt ttaaccaggg
     5221 ctgcgccctg tgcgcgtgac cgcgcacgcc gaaggggggt gccccccctt ctcgaaccct
     5281 cccggcccgc taacgcgggc ctcccatccc cccaggggct gcgcccctcg gccgcgaacg
     5341 gcctcacccc aaaaatggca gcgctggcag tccttgccat tgccgggatc ggggcagtaa
     5401 cgggatgggc gatcagcccg aaagctaccc ctatttgttt atttttctaa atacattcaa
     5461 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     5521 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     5581 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     5641 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     5701 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     5761 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5821 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     5881 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     5941 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     6001 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     6061 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     6121 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     6181 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     6241 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     6301 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     6361 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     6421 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     6481 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     6541 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     6601 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     6661 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     6721 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     6781 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     6841 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     6901 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     6961 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     7021 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     7081 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     7141 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     7201 acatggactc tagctagagg atcacaggca gcaacgctct gtcatcgtta caatcaacat
     7261 gctaccctcc gcgagatcat ccgtgtttca aacccggcag cttagttgcc gttcttccga
     7321 atagcatcgg taacatgagc aaagtctgcc gccttacaac ggctctcccg ctgacgccgt
     7381 cccggactga tgggctgcct gtatcgagtg gtgattttgt gccgagctgc cggtcgggga
     7441 gctgttggct ggctggtggc aggatatatt gtgg
//
