LOCUS       sequence-191806-        6930 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            MAP3K10-1/1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6930)
  AUTHORS   Teyra J, Huang H, Jain S, Guan X, Dong A, Liu Y, Tempel W, Min J, 
            Tong Y, Kim PM, Bader GD, Sidhu SS
  TITLE     Comprehensive Analysis of the Human SH3 Domain Family Reveals a Wide
            Variety of Non-canonical Specificities.
  JOURNAL   Structure. 2017 Oct 3;25(10):1598-1610.e3. doi: 
            10.1016/j.str.2017.07.017. Epub 2017 Sep 7.
  PUBMED    28890361
REFERENCE   2  (bases 1 to 6930)
  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: 
            "10.1016/j.str.2017.07.017"; journalName: "Structure"; date: 
            "2017-10-3- 3"; volume: "25"; issue: "10"; pages: "1598-1610.e3"
COMMENT     Sequence Label: pHH0103_MAP3K10-1/1
FEATURES             Location/Qualifiers
     source          1..6930
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     25..43
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        66..143
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             144..1226
                     /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     complement(163..182)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1469..1474,1475..1490,1491..1497)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1505..1521
                     /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)."
     primer_bind     1510..1532
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1606..1623
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1639..2292
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2250..2272
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2299..2322
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2335..2352
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2593..2640
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4233..4688)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4370..4391)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4582..4601
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4730..4752)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4838..4978
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4893..4912
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4993..5010)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5164..5751)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5922..6713,6714..6782))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6545..6564
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6783..6887)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg tcttcaagaa
       61 ttcccgacac catcgaatgg tgcaaaacct ttcgcggtat ggcatgatag cgcccggaag
      121 agagtcaatt cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc gcagagtatg
      181 ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac gtttctgcga
      241 aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg
      301 cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc agtctggccc
      361 tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca
      421 gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca
      481 atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat gaccaggatg
      541 ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat gtctctgacc
      601 agacacccat caacagtatt attttctccc atgaagacgg tacgcgactg ggcgtggagc
      661 atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggcccatta agttctgtct
      721 cggcgcgtct gcgtctggct ggctggcata aatatctcac tcgcaatcaa attcagccga
      781 tagcggaacg ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc
      841 tgaatgaggg catcgttccc actgcgatgc tggttgccaa cgatcagatg gcgctgggcg
      901 caatgcgcgc cattaccgag tccgggctgc gcgttggtgc ggatatctcg gtagtgggat
      961 acgacgatac cgaagacagc tcatgttata tcccgccgtt aaccaccatc aaacaggatt
     1021 ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca actctctcag ggccaggcgg
     1081 tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc ctggcgccca
     1141 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
     1201 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtaagtta gctcactcat
     1261 taggcacaat tctcatgttt gacagcttat catcgactgc acggtgcacc aatgcttctg
     1321 gcgtcaggca gccatcggaa gctgtggtat ggctgtgcag gtcgtaaatc actgcataat
     1381 tcgtgtcgct caaggcgcac tcccgttctg gataatgttt tttgcgccga catcataacg
     1441 gttctggcaa atattctgaa atgagctgtt gacaattaat catcggctcg tataatgtgt
     1501 ggaattgtga gcggataaca atttcacaca ggaaacagcc agtccgttta ggtgttttca
     1561 cgagcacttc accaacaagg accatagatt atgaaaatcg aagaacacca tcaccatcac
     1621 cattccagcg gtaagcttat gtcccctata ctaggttatt ggaaaattaa gggccttgtg
     1681 caacccactc gacttctttt ggaatatctt gaagaaaaat atgaagagca tttgtatgag
     1741 cgcgatgaag gtgataaatg gcgaaacaaa aagtttgaat tgggtttgga gtttcccaat
     1801 cttccttatt atattgatgg tgatgttaaa ttaacacagt ctatggccat catacgttat
     1861 atagctgaca agcacaacat gttgggtggt tgtccaaaag agcgtgcaga gatttcaatg
     1921 cttgaaggag cggttttgga tattagatac ggtgtttcga gaattgcata tagtaaagac
     1981 tttgaaactc tcaaagttga ttttcttagc aagctacctg aaatgctgaa aatgttcgaa
     2041 gatcgtttat gtcataaaac atatttaaat ggtgatcatg taacccatcc tgacttcatg
     2101 ttgtatgacg ctcttgatgt tgttttatac atggacccaa tgtgcctgga tgcgttccca
     2161 aaattagttt gttttaaaaa acgtattgaa gctatcccac aaattgataa gtacttgaaa
     2221 tccagcaagt atatagcatg gcctttgcag ggctggcaag ccacgtttgg tggtggcgac
     2281 catcctccaa aatcggatct ggaagttctg ttccaggggc ccctgtccag cggtctggtt
     2341 ccgcgtggtt ccggtaccgc ggcccagccg gccatggccc agggcgcgct gccagtttgg
     2401 accgccgttt tcgattatga agccgccggt gatgaagaac tgaccctgcg tcgtggtgat
     2461 cgtgttcaag ttctgtccca ggatcgtgca gtttctggtg atgaaggatg gtggaccggt
     2521 caattaccat ctggtcgtgt tggtgttttt ccgtccaact atgttgcacc aggtgcagcg
     2581 gccgcataat aaaccgatac aattaaaggc tccttttgga gccttttttt tggagatttt
     2641 caacgtgaaa aaattattat tcgcaattcc tttagttgtt cctttctatt ctcactccgc
     2701 tgaaactgtt gaaagttgtt tagcaaaacc ccatacagaa aattcattta ctaacgtctg
     2761 gaaagacgac aaaactttag atcgttacgc taactatgag ggctgtctgt ggaatgctac
     2821 aggcgttgta gtttgtactg gtgacgaaac tcagtgtcta gctagagtgg cggtggctct
     2881 ggttccggtg attttgatta tgaaaagatg gcaaacgcta ataagggggc tatgaccgaa
     2941 aatgccgatg aaaacgcgct acagtctgac gctaaaggca aacttgattc tgtcgctact
     3001 gattacggtg ctgctatcga tggtttcatt ggtgacgttt ccggccttgc taatggtaat
     3061 ggtgctactg gtgattttgc tggctctaat tcccaaatgg ctcaagtcgg tgacggtgat
     3121 aattcacctt taatgaataa tttccgtcaa tatttacctt ccctccctca atcggttgaa
     3181 tgtcgccctt ttgtctttgg cgctggtaaa ccatatgaat tttctattga ttgtgacaaa
     3241 ataaacttat tccgtggtgt ctttgcgttt cttttatatg ttgccacctt tatgtatgta
     3301 ttttctacgt ttgctaacat actgcgtaat aaggagtctt aatcatgcca gttcttttgg
     3361 ctagcgccgc cctatacctt gtctgcctcc ccgcgttgcg tcgcggtgca tggagccggg
     3421 ccacctcgac ctgaatggaa gccggcggca cctcgctaac ggattcacca ctccaagaat
     3481 tggagccaat caattcttgc ggagaactgt gaatgcgcaa accaaccctt ggcagaacat
     3541 atccatcgcg tccgccatct ccagcagccg cacgcggcgc atctcgggca gcgttgggtc
     3601 ctggccacgg gtgcgcatga tcgtgctcct gtcgttgagg acccggctag gctggcgggg
     3661 ttgccttact ggttagcaga atgaatcacc gatacgcgag cgaacgtgaa gcgactgctg
     3721 ctgcaaaacg tctgcgacct gagcaacaac atgaatggtc ttcggtttcc gtgtttcgta
     3781 aagtctggaa acgcggaagt cagcgccctg caccattatg ttccggatct gcatcgcagg
     3841 atgctgctgg ctaccctgtg gaacacctac atctgtatta acgaagcgct ggcattgacc
     3901 ctgagtgatt tttctctggt cccgccgcat ccataccgcc agttgtttac cctcacaacg
     3961 ttccagtaac cgggcatgtt catcatcagt aacccgtatc gtgagcatcc tctctcgttt
     4021 catcggtatc attaccccca tgaacagaaa tcccccttac acggaggcat cagtgaccaa
     4081 acaggaaaaa accgccctta acatggcccg ctttatcaga agccagacat taacgcttct
     4141 ggagaaactc aacgagctgg acgcggatga acaggcagac atctgtgaat cgcttcacga
     4201 ccacgctgat gagctttacc gcaggatccg cgaaattgta agcgttaata ttttgttaaa
     4261 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     4321 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     4381 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     4441 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     4501 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     4561 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     4621 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     4681 gggcgcgtcg cggatcctgc ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca
     4741 tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc
     4801 gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc cagtcacgta
     4861 gcgatagcgg agtgtatact ggcttaacta tgcggcatca gagcagattg tactgagagt
     4921 gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg
     4981 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     5041 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     5101 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     5161 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     5221 gtggcgaaac ccgacaggac tataaagata ccagggtttc cccctggaag ctccctcgtg
     5281 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     5341 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     5401 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     5461 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     5521 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     5581 cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt
     5641 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     5701 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     5761 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     5821 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     5881 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     5941 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     6001 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     6061 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     6121 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     6181 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctgca
     6241 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     6301 tcaaggcgag ttacatgatc ccccatattg tgcaaaaaag cggttagctc cttcggtcct
     6361 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     6421 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     6481 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaaca
     6541 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     6601 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     6661 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     6721 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     6781 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     6841 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     6901 aaagtgccac ctgacgtcta agaaaccatt
//
