LOCUS       sequence-350012-        6715 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of SnRK2.3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6715)
  AUTHORS   Ng LM, Soon FF, Zhou XE, West GM, Kovach A, Suino-Powell KM, 
            Chalmers MJ, Li J, Yong EL, Zhu JK, Griffin PR, Melcher K, Xu HE
  TITLE     Structural basis for basal activity and autoactivation of abscisic 
            acid (ABA) signaling SnRK2 kinases.
  JOURNAL   Proc Natl Acad Sci U S A. 2011 Dec 27;108(52):21259-64. doi: 
            10.1073/pnas.1118651109. Epub 2011 Dec 12.
  PUBMED    22160701
REFERENCE   2  (bases 1 to 6715)
  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: 
            "10.1073/pnas.1118651109"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2011-12-27- 27"; volume: "108"; issue: "52"; pages: 
            "21259-64"
COMMENT     Sequence Label: pSUMO His6 SUMO SnRK2.3 (19-361)
FEATURES             Location/Qualifiers
     source          1..6715
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     956..975
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(995..1072)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1278..1297
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1381..1400
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1381..1399
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1400..1424
                     /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             1439..1461
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1474..1491
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1495..1785
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="SDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPLR
                     RLMEAFAKRQGKEMDSLRFLYDGIRIQADQAPEDLDMEDNDIIEAHREQIGG"
     CDS             2827..2844
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2897..2915)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2911..2958
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2995..3450
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3082..3101)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3292..3313
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3476..3580
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3581..3649,3650..4441)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3799..3818)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4612..5200
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5101..5120
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5354..5371
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5386..5525
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5451..5470)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5611..5633
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5627..5818)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6627..6715,1..994))
                     /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"
ORIGIN
        1 gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg
       61 cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat
      121 cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc
      181 ccactaccga gatatccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc
      241 ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca
      301 tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg
      361 gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga
      421 cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct
      481 ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt
      541 cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat
      601 cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt
      661 gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg
      721 cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg
      781 ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca
      841 cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg
      901 cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat
      961 actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt
     1021 ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct
     1081 cgacgctctc ccttatgcga ctcctgcatt aggaagcagc ccagtagtag gttgaggccg
     1141 ttgagcaccg ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg
     1201 gccacggggc ctgccaccat acccacgccg aaacaagcgc tcatgagccc gaagtggcga
     1261 gcccgatctt ccccatcggt gatgtcggcg atataggcgc cagcaaccgc acctgtggcg
     1321 ccggtgatgc cggccacgat gcgtccggcg tagaggatcg agatctcgat cccgcgaaat
     1381 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
     1441 tgtttaactt taagaaggag atataccatg ggtcatcacc atcatcatca cgggtcggac
     1501 tcagaagtca atcaagaagc taagccagag gtcaagccag aagtcaagcc tgagactcac
     1561 atcaatttaa aggtgtccga tggatcttca gagatcttct tcaagatcaa aaagaccact
     1621 cctttaagaa ggctgatgga agcgttcgct aaaagacagg gtaaggaaat ggactcctta
     1681 agattcttgt acgacggtat tagaattcaa gctgatcagg cccctgaaga tttggacatg
     1741 gaggataacg atattattga ggctcaccgc gaacagattg gaggtggaag tgatcgatat
     1801 gacttcgtta aggatattgg ttctggtaat ttcggtgttg ctcgtcttat gagagataaa
     1861 ctcactaaag agcttgttgc tgtcaagtac atcgagagag gagacaagat tgatgaaaat
     1921 gttcaaaggg agatcattaa ccacaggtca ctaaggcatc ctaatattgt cagatttaaa
     1981 gaggtcattt tgacgccgac tcatctggct atcataatgg aatatgcttc tggcggtgaa
     2041 ctttacgagc ggatttgcaa tgcaggacgg tttagtgaag atgaggctcg gttcttcttt
     2101 cagcagcttc tatctggagt cagttattgt cattcgatgc aaatttgcca tcgtgacctg
     2161 aagctagaga atacattgtt ggatggaagt cctgctcctc gattaaaaat ttgtgatttt
     2221 ggatattcaa agtcttctgt tcttcattca caaccaaagt caactgttgg tactcctgca
     2281 tacatcgctc cagaggtact gcttcgtcag gaatatgatg gcaagattgc agatgtatgg
     2341 tcatgtggtg tgaccttata cgtcatgttg gttggagcgt atccgttcga agatccagaa
     2401 gagccaagag actatcggaa aacaatacag agaatcctta gcgttaaata ctcaatccct
     2461 gatgacatac ggatatcacc tgaatgctgt catcttattt caagaatctt cgtggctgat
     2521 cccgctacca gaataagcat accagagatc aaaacccata gttggttctt gaagaatctc
     2581 cctgctgatc taatgaacga gagcaacaca ggaagccagt tccaggagcc tgaacaacca
     2641 atgcaaagcc ttgacacaat catgcaaatc atctctgaag ccacaattcc cgctgttcga
     2701 aaccgttgcc tagacgattt catgactgac aatcttgatc ttgacgatga catggatgac
     2761 tttgactctg aatctgaaat cgacattgac agtagcggag agatagttta cgctctctaa
     2821 ctcgagcacc accaccacca ccactgagat ccggctgcta acaaagcccg aaaggaagct
     2881 gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg
     2941 gtcttgaggg gttttttgct gaaaggagga actatatccg gattggcgaa tgggacgcgc
     3001 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     3061 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     3121 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     3181 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     3241 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     3301 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     3361 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     3421 attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg
     3481 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     3541 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     3601 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     3661 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     3721 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     3781 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     3841 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     3901 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     3961 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     4021 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     4081 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgcagcaa tggcaacaac
     4141 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     4201 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     4261 gtttattgct gataaatctg gagccggtga gcgtggttct cgcggtatca ttgcagcact
     4321 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     4381 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     4441 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     4501 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     4561 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     4621 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     4681 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     4741 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     4801 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     4861 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     4921 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     4981 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     5041 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     5101 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     5161 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     5221 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     5281 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     5341 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
     5401 tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag tacaatctgc
     5461 tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg
     5521 ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     5581 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac
     5641 cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg
     5701 attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta
     5761 atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt ttggtcactg
     5821 atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg aaacgagaga
     5881 ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa cgttgtgagg
     5941 gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag ggtcaatgcc
     6001 agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat cctgcgatgc
     6061 agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt tacgaaacac
     6121 ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc
     6181 ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa ccccgccagc
     6241 ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggggcc gccatgccgg
     6301 cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg aaggcttgag
     6361 cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc
     6421 gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct acgagttgca
     6481 tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg
     6541 agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga
     6601 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     6661 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgg
//
