LOCUS       sequence-350011-        6499 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of D14.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6499)
  AUTHORS   Zhao LH, Zhou XE, Wu ZS, Yi W, Xu Y, Li S, Xu TH, Liu Y, Chen RZ, 
            Kovach A, Kang Y, Hou L, He Y, Xie C, Song W, Zhong D, Xu Y, Wang Y,
            Li J, Zhang C, Melcher K, Xu HE
  TITLE     Crystal structures of two phytohormone signal-transducing alpha/beta
            hydrolases: karrikin-signaling KAI2 and strigolactone-signaling 
            DWARF14.
  JOURNAL   Cell Res. 2013 Mar;23(3):436-9. doi: 10.1038/cr.2013.19. Epub 2013 
            Feb 5.
  PUBMED    23381136
REFERENCE   2  (bases 1 to 6499)
  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: 
            "10.1038/cr.2013.19"; journalName: "Cell Res"; date: "2013-03"; 
            volume: "23"; issue: "3"; pages: "436-9"
COMMENT     Sequence Label: pET Duet1 His6 SUMO D14
FEATURES             Location/Qualifiers
     source          1..6499
                     /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             2611..2628
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2681..2699)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2695..2742
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2779..3234
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2866..2885)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3076..3097
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3260..3364
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3365..3433,3434..4225)
                     /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(3583..3602)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4396..4984
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4885..4904
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5138..5155
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5170..5309
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5235..5254)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5395..5417
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5411..5602)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6411..6499,1..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 gaggtggatc ctcgggcgca
     1801 agtctgctgc aaatcctgaa tgtccgtgtg gtcggctctg gtgaacgtgt tgtcgtcctg
     1861 tcgcatggct tcggtacgga tcagagtgcg tggtcccgtg tgctgccgta tctgacccgt
     1921 gatcatcgcg tggttctgta cgacctggtt tgcgctggca gcgtcaaccc ggatcacttt
     1981 gacttccgtc gctatgataa tctggacgca tacgttgatg acctgctggc tattctggat
     2041 gcgctgcgta tcccgcgctg tgcctttgtt ggccatagcg tctctgccat gattggtatc
     2101 ctggcaagca ttcgtcgccc ggacctgttt gctaaactgg tgctgatcgg tgcgagcccg
     2161 cgcttcctga acgattctga ctatcacggc ggttttgaac tggaagaaat tcagcaagtt
     2221 ttcgatgcaa tgggtgctaa ttattctgca tgggcaaccg gttacgcacc gctggccgtg
     2281 ggtgcagacg ttccggcggc cgtccaggaa tttagtcgta cgctgttcaa catgcgcccg
     2341 gatatctccc tgcatgtgtg ccaaaccgtt tttaaaacgg acctgcgtgg tgtcctgggt
     2401 atggtgcgtg caccgtgtgt cgtggttcag accacgcgtg atgtctcagt gccggcctcg
     2461 gtggcagctt acctgaaagc acatctgggc ggtcgcacca cggttgaatt tctgcaaacc
     2521 gaaggtcacc tgccgcatct gtctgccccg tcactgctgg ctcaggtgct gcgtcgtgct
     2581 ctggcccgtt actaagcggc cgcactcgag caccaccacc accaccactg agatccggct
     2641 gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca
     2701 taaccccttg gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata
     2761 tccggattgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     2821 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     2881 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     2941 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     3001 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     3061 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     3121 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     3181 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg
     3241 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     3301 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     3361 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     3421 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     3481 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     3541 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     3601 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     3661 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     3721 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     3781 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     3841 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     3901 gatgcctgca gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     3961 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     4021 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     4081 ttctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     4141 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     4201 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     4261 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     4321 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     4381 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     4441 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     4501 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
     4561 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     4621 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     4681 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     4741 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     4801 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     4861 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     4921 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     4981 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     5041 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     5101 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     5161 ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcaa
     5221 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc
     5281 tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc
     5341 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     5401 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa
     5461 gctcatcagc gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct
     5521 cgttgagttt ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg
     5581 cggttttttc ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg
     5641 taatgatacc gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg
     5701 cccggttact ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga
     5761 gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg
     5821 gtagccagca gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc
     5881 gcgtttccag actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg
     5941 cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct
     6001 aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc
     6061 gcacccgtgg ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg
     6121 cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca
     6181 ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg
     6241 ccggcacctg tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag
     6301 tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc
     6361 gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg
     6421 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     6481 gaggcggttt gcgtattgg
//
