LOCUS       sequence-350025-        6742 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of SnRK2.6.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6742)
  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 6742)
  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 SnRK2.6 (11-362)
FEATURES             Location/Qualifiers
     source          1..6742
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    13..37
                     /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             52..74
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             87..104
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             108..398
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="SDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPLR
                     RLMEAFAKRQGKEMDSLRFLYDGIRIQADQAPEDLDMEDNDIIEAHREQIGG"
     CDS             1467..1484
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1537..1555)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1551..1598
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1635..2090
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1722..1741)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1932..1953
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2116..2220
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2221..2289,2290..3081)
                     /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(2439..2458)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3252..3840
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3741..3760
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3994..4011
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4026..4165
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4091..4110)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4251..4273
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4267..4458)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5267..6349)
                     /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     6311..6330
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6350..6427)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6633..6652
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(6736..6742,1..13)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(6736..6742,1..12)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 actcactata ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa
       61 ctttaagaag gagatatacc atgggtcatc accatcatca tcacgggtcg gactcagaag
      121 tcaatcaaga agctaagcca gaggtcaagc cagaagtcaa gcctgagact cacatcaatt
      181 taaaggtgtc cgatggatct tcagagatct tcttcaagat caaaaagacc actcctttaa
      241 gaaggctgat ggaagcgttc gctaaaagac agggtaagga aatggactcc ttaagattct
      301 tgtacgacgg tattagaatt caagctgatc aggcccctga agatttggac atggaggata
      361 acgatattat tgaggctcac cgcgaacaga ttggaggtgg agatttgccg attatgcacg
      421 atagtgatag gtatgaactc gtcaaggata ttggctccgg taattttgga gttgcgagat
      481 tgatgagaga caagcaaagt aatgagcttg ttgctgttaa atatatcgag agaggtgaga
      541 agatagatga aaatgtaaaa agggagataa tcaaccacag gtccttaaga catcccaata
      601 tcgttagatt caaagaggtt atattaacac caacccattt agccattgtt atggaatatg
      661 catctggagg agaacttttc gagcgaatct gcaatgcagg ccgcttcagc gaagacgagg
      721 cgaggttttt cttccagcaa ctcatttcag gagttagtta ctgtcatgct atgcaagtat
      781 gtcaccgaga cttaaagctc gagaatacgt tattagatgg tagcccggcc cctcgtctaa
      841 agatatgtga tttcggatat tctaagtcat cagtgttaca ttcgcaacca aaatcaactg
      901 ttggaactcc tgcttacatc gctcctgagg ttttactaaa gaaagaatat gatggaaagg
      961 ttgcagatgt ttggtcttgt ggggttactc tgtatgtcat gctggttgga gcatatcctt
     1021 tcgaagatcc cgaggaacca aagaatttca ggaaaactat acatagaatc ctgaatgttc
     1081 agtatgctat tccggattat gttcacatat ctcctgaatg tcgccatttg atctccagaa
     1141 tatttgttgc tgaccctgca aagaggatat caattcctga aataaggaac catgaatggt
     1201 ttctaaagaa tctaccggca gatctaatga acgataacac gatgaccact cagtttgatg
     1261 aatcggatca accgggccaa agcatagaag aaattatgca gatcattgca gaagcaactg
     1321 ttcctcctgc aggcacccag aatctgaacc attacctcac aggaagcttg gacatagatg
     1381 acgatatgga ggaagactta gagagcgacc ttgatgatct tgacatcgac agtagcggag
     1441 agattgtgta cgcaatgtga gtcgagcacc accaccacca ccactgagat ccggctgcta
     1501 acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac
     1561 cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg
     1621 gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     1681 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     1741 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     1801 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     1861 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     1921 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     1981 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     2041 ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact
     2101 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     2161 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     2221 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     2281 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     2341 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     2401 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     2461 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     2521 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     2581 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     2641 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     2701 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     2761 cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     2821 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     2881 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtggttct
     2941 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     3001 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     3061 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     3121 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     3181 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     3241 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     3301 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     3361 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     3421 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     3481 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     3541 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     3601 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     3661 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     3721 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     3781 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     3841 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     3901 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     3961 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     4021 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcaatggt
     4081 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc
     4141 gctacgtgac tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg
     4201 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     4261 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc
     4321 atcagcgtgg tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt
     4381 gagtttctcc agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt
     4441 tttttcctgt ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat
     4501 gataccgatg aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg
     4561 gttactggaa cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa
     4621 aatcactcag ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag
     4681 ccagcagcat cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt
     4741 ttccagactt tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga
     4801 cgttttgcag cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc
     4861 agtaaggcaa ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac
     4921 ccgtggggcc gccatgccgg cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg
     4981 accagtgacg aaggcttgag cgagggcgtg caagattccg aataccgcaa gcgacaggcc
     5041 gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg
     5101 cacctgtcct acgagttgca tgataaagaa gacagtcata agtgcggcga cgatagtcat
     5161 gccccgcgcc caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga
     5221 tcccggtgcc taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt
     5281 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     5341 cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct
     5401 gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc
     5461 ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt
     5521 cggtatcgtc gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa
     5581 tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga
     5641 tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt
     5701 cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac
     5761 gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca
     5821 atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt
     5881 tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt
     5941 ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt
     6001 gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg
     6061 acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg
     6121 acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg
     6181 ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt
     6241 tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat
     6301 aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc
     6361 tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga
     6421 tggtgtccgg gatctcgacg ctctccctta tgcgactcct gcattaggaa gcagcccagt
     6481 agtaggttga ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg
     6541 cccaacagtc ccccggccac ggggcctgcc accataccca cgccgaaaca agcgctcatg
     6601 agcccgaagt ggcgagcccg atcttcccca tcggtgatgt cggcgatata ggcgccagca
     6661 accgcacctg tggcgccggt gatgccggcc acgatgcgtc cggcgtagag gatcgagatc
     6721 tcgatcccgc gaaattaata cg
//
