LOCUS       sequence-465271-        6243 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Recombinant expression of His6-tagged Arabidopsis thaliana MKK5 in 
            E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6243)
  AUTHORS   Simranjit Kaur, Thomas Colby, Domenika Thieme, Carsten Proksch, 
            Susanne Matschi, eIvan Matic,, Lennart Wirthmueller
  TITLE     Untargeted proteomics identifies plant substrates of the 
            bacterial-derived ADP-ribosyltransferase AvrRpm1
  JOURNAL   bioRxiv 2023.09.25.558804
REFERENCE   2  (bases 1 to 6243)
  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: "bioRxiv 
            2023.09.25.558804"
COMMENT     Sequence Label: pOPINF AtMKK5
FEATURES             Location/Qualifiers
     source          1..6243
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_recomb     181..1008
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     CDS             181..305
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(343..948)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     enhancer        1140..1443
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1449..1726
                     /label=chicken beta-actin promoter
     primer_bind     2005..2027
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2089..2108
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     2150..2169
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2150..2168
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2169..2193
                     /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        2209..2318
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     CDS             2339..2356
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2366..2389
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(3538..3557)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3603..3658
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3657..3676)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      3746..3793
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     3806..4511
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           3894..4266
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(3923..4511)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    complement(4520..4553)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(4721..5308)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4800..4819)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5321..6112,6113..6181))
                     /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     5944..5963
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tcctgcatct tttaatcaaa tcccaagatg tgtataaacg cgccggtatg tacaggaaga
       61 ggtttatact aaactgttac attgcaaacg tggtttcgtg tgccaagtgt gaaaaccgat
      121 gtttaatcaa ggctctgacg catttctaca accacgactc caagtgtgtg ggtgaagtca
      181 tgcatctttt aatcaaatcc caagatgtgt ataaaccacc aaactgccaa aaaatgaaaa
      241 ctgtcgacaa gctctgtccg tttgctggca actgcaaggg tctcaatcct atttgtaatt
      301 attgaataat aaaacaatta taaatgtcaa atttgttttt tattaacgat acaaaccaaa
      361 cgcaacaaga acatttgtag tattatctat aattgaaaac gcgtagttat aatcgctgag
      421 gtaatattta aaatcatttt caaatgattc acagttaatt tgcgacaata taattttatt
      481 ttcacataaa ctagacgcct tgtcgtcttc ttcttcgtat tccttctctt tttcattttt
      541 ctcttcataa aaattaacat agttattatc gtatccatat atgtatctat cgtatagagt
      601 aaattttttg ttgtcataaa tatatatgtc ttttttaatg gggtgtatag taccgctgcg
      661 catagttttt ctgtaattta caacagtgct attttctggt agttcttcgg agtgtgttgc
      721 tttaattatt aaatttatat aatcaatgaa tttgggatcg tcggttttgt acaatatgtt
      781 gccggcatag tacgcagctt cttctagttc aattacacca ttttttagca gcaccggatt
      841 aacataactt tccaaaatgt tgtacgaacc gttaaacaaa aacagttcac ctcccttttc
      901 tatactattg tctgcgagca gttgtttgtt gttaaaaata acagccattg taatgagacg
      961 cacaaactaa tatcacaaac tggaaatgtc tatcaatata tagttgctga tggccggccg
     1021 taatgagacg cacaaactaa tatcacaaac tggaaatgtc tatcaatata tagttgctct
     1081 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
     1141 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
     1201 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
     1261 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
     1321 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
     1381 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
     1441 atgcatggtc gaggtgagcc ccacgttctg cttcactctc cccatctccc ccccctcccc
     1501 acccccaatt ttgtatttat ttatttttta attattttgt gcagcgatgg gggcgggggg
     1561 gggggggggg cgcgcgccag gcggggcggg gcggggcgag gggcggggcg gggcgaggcg
     1621 gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag
     1681 gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg
     1741 ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact
     1801 gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctccttcgg gctgtaatta
     1861 gcgcttggtt taatgacggc ttgtttcttt tctgtggctg cgtgaaagcc ttgaggggct
     1921 ccgggagggc cctttgtgcg gggggagcgg ctcggggctg tccgcggggg gacggctgcc
     1981 ttcggggggg acggggcagg gcggggttcg gcttctggcg tgtgaccggc ggctctagag
     2041 cctctgctaa ccatgttcat gccttcttct ttttcctaca gctcctgggc aacgtgctgg
     2101 ttattgtgct gtctcatcat tttggcaaag aattggatcg gaccgaaatt aatacgactc
     2161 actatagggg aattgtgagc ggataacaat tccccggagt taatccggga cctttaattc
     2221 aacccaacac aatatattat agttaaataa gaattattat caaatcattt gtatattaat
     2281 taaaatacta tactgtaaat tacattttat ttacaatcaa aggagatata ccatggcaca
     2341 ccatcaccac catcacagca gcggtctgga agttctgttt cagggcccga aaccgattca
     2401 atctccttct ggagtagctt cacctatgaa gaaccgttta cgcaaacgtc ctgacctaag
     2461 cttaccactc ccacaccgcg acgtcgctct cgccgtacct ctccctctcc cacctccttc
     2521 ttcctcttca tccgctccgg cgtcttcctc cgcgatctca accaacatct ccgccgctaa
     2581 aagcttatcc gagctagaac gagtgaaccg aatcggaagc ggagccggag gaacggttta
     2641 caaagtaatc cacactccga cgtcacgtcc tttcgctctc aaagtgattt acggaaacca
     2701 cgaagatacc gtgagacgtc agatctgtag agagatcgag atcttaagaa gtgttgatca
     2761 tccaaacgtt gtgaaatgtc acgatatgtt tgatcataac ggtgagatcc aggttttgct
     2821 tgagtttatg gatcaaggat ctcttgaagg agctcatata tggcaagaac aggaattagc
     2881 tgatctctct cgtcagattc ttagtggatt agcttatctt catcgtcgtc atatcgttca
     2941 tcgtgatatc aaaccttcga atcttcttat aaactcagct aaaaatgtga aaattgctga
     3001 ttttggtgtg agtaggatct tggcacaaac aatggatcct tgtaattcat ctgttggtac
     3061 tattgcttat atgagtcctg agaggattaa tactgatttg aatcatggtc gttacgatgg
     3121 ttatgctgga gatgtttgga gtttaggtgt tagtatcttg gagttttact tggggaggtt
     3181 tccttttgct gtgagtagac aaggtgattg ggctagtctt atgtgtgcta tttgtatgtc
     3241 tcagccacct gaagctccgg ctacggcgtc tcaggagttt cgtcactttg tttcttgttg
     3301 tttacagagt gatcctccta agagatggtc agctcaacag cttttgcagc atcctttcat
     3361 acttaaagct accggtggtc ctaatctccg tcaaatgttg ccgccgcctc gtcctcttcc
     3421 ttctgcctct tgaagctttc tagaccagtt tgtgattaac ctcaggtgca ggctgcctat
     3481 cagaaggtgg tggctggtgt ggccaatgcc ctggctcaca aataccactg agatcgatct
     3541 ttttccctct gccaaaaatt atggggacat catgaagccc cttgagcatc tgacttctgg
     3601 ctaataaagg aaatttattt tcattgcaat agtgtgttgg aattttttgt gtctctcact
     3661 cggaaggaca tatgggaggg caaatcattt aaaacatcag aatgagtatt tggtttagag
     3721 tttggcaaca tatgcccata tgtaactagc ataacccctt ggggcctcta aacgggtctt
     3781 gaggggtttt ttgctgaaag catgcggagg aaattctcct tgaagtttcc ctggtgttca
     3841 aagtaaagga gtttgcacca gacgcacctc tgttcactgg tccggcgtat taaaacacga
     3901 tacattgtta ttagtacatt tattaagcgc tagattctgt gcgttgttga tttacagaca
     3961 attgttgtac gtattttaat aattcattaa atttataatc tttagggtgg tatgttagag
     4021 cgaaaatcaa atgattttca gcgtctttat atctgaattt aaatattaaa tcctcaatag
     4081 atttgtaaaa taggtttcga ttagtttcaa acaagggttg tttttccgaa ccgatggctg
     4141 gactatctaa tggattttcg ctcaacgcca caaaacttgc caaatcttgt agcagcaatc
     4201 tagctttgtc gatattcgtt tgtgttttgt tttgtaataa aggttcgacg tcgttcaaaa
     4261 tattatgcgc ttttgtattt ctttcatcac tgtcgttagt gtacaattga ctcgacgtaa
     4321 acacgttaaa tagagcttgg acatatttaa catcgggcgt gttagcttta ttaggccgat
     4381 tatcgtcgtc gtcccaaccc tcgtcgttag aagttgcttc cgaagacgat tttgccatag
     4441 ccacacgacg cctattaatt gtgtcggcta acacgtccgc gatcaaattt gtagttgagc
     4501 tttttggaat tgcgatcgca taacttcgta tagcatacat tatacgaagt tataagctcg
     4561 gaacgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4621 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     4681 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     4741 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     4801 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     4861 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     4921 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     4981 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     5041 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     5101 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     5161 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     5221 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     5281 gattacgcgc agaaaaaaag gatctcaaga agatcctttg ttaccaatgc ttaatcagtg
     5341 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     5401 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     5461 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     5521 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     5581 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     5641 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     5701 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     5761 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     5821 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     5881 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     5941 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6001 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     6061 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     6121 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     6181 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgtccgcgc
     6241 gtt
//
