LOCUS       sequence-191778-        7002 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            MAP3K9-1/2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7002)
  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 7002)
  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_MAP3K9-1/2
FEATURES             Location/Qualifiers
     source          1..7002
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     12..30
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        53..130
                     /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             131..1213
                     /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(150..169)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1456..1461,1462..1477,1478..1484)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1492..1508
                     /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     1497..1519
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1593..1610
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1626..2279
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2237..2259
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2286..2309
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2322..2339
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2652..2699
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4292..4747)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4429..4450)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4641..4660
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4789..4811)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4897..5037
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4952..4971
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5052..5069)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5223..5810)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5981..6772,6773..6841))
                     /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     6604..6623
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6842..6946)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 taacctataa aaataggcgt atcacgaggc cctttcgtct tcaagaattc ccgacaccat
       61 cgaatggtgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga gtcaattcag
      121 ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta
      181 tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa
      241 agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc
      301 gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc
      361 gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc
      421 gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca
      481 acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga
      541 agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa
      601 cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt
      661 gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg
      721 tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga
      781 aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat
      841 cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat
      901 taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg acgataccga
      961 agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg
     1021 gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca
     1081 gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc
     1141 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     1201 aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag gcacaattct
     1261 catgtttgac agcttatcat cgactgcacg gtgcaccaat gcttctggcg tcaggcagcc
     1321 atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg tgtcgctcaa
     1381 ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt ctggcaaata
     1441 ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga attgtgagcg
     1501 gataacaatt tcacacagga aacagccagt ccgtttaggt gttttcacga gcacttcacc
     1561 aacaaggacc atagattatg aaaatcgaag aacaccatca ccatcaccat tccagcggta
     1621 agcttatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa cccactcgac
     1681 ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc gatgaaggtg
     1741 ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt ccttattata
     1801 ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata gctgacaagc
     1861 acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt gaaggagcgg
     1921 ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt gaaactctca
     1981 aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat cgtttatgtc
     2041 ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg tatgacgctc
     2101 ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa ttagtttgtt
     2161 ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc agcaagtata
     2221 tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat cctccaaaat
     2281 cggatctgga agttctgttc caggggcccc tgtccagcgg tctggttccg cgtggttccg
     2341 gtaccgcggc ccagccggcc gaagaagaag aagaagccgc tgctgctgtt ggtccgggtg
     2401 aactgggttg cgatgctccg ctgccgtatt ggaccgctgt ttttgaatat gaagcggccg
     2461 gcgaagatga actgaccctg cgtctgggtg acgtggttga agtcctgagt aaagattccc
     2521 aggtgtcagg cgacgaaggt tggtggaccg gccagctgaa ccaacgcgtt ggtatttttc
     2581 cgtcgaatta cgtcacgccg cgttctgcgt tcagctctcg ctgccaaccg ggcggtgcgg
     2641 ccgcataata aaccgataca attaaaggct ccttttggag cctttttttt ggagattttc
     2701 aacgtgaaaa aattattatt cgcaattcct ttagttgttc ctttctattc tcactccgct
     2761 gaaactgttg aaagttgttt agcaaaaccc catacagaaa attcatttac taacgtctgg
     2821 aaagacgaca aaactttaga tcgttacgct aactatgagg gctgtctgtg gaatgctaca
     2881 ggcgttgtag tttgtactgg tgacgaaact cagtgtctag ctagagtggc ggtggctctg
     2941 gttccggtga ttttgattat gaaaagatgg caaacgctaa taagggggct atgaccgaaa
     3001 atgccgatga aaacgcgcta cagtctgacg ctaaaggcaa acttgattct gtcgctactg
     3061 attacggtgc tgctatcgat ggtttcattg gtgacgtttc cggccttgct aatggtaatg
     3121 gtgctactgg tgattttgct ggctctaatt cccaaatggc tcaagtcggt gacggtgata
     3181 attcaccttt aatgaataat ttccgtcaat atttaccttc cctccctcaa tcggttgaat
     3241 gtcgcccttt tgtctttggc gctggtaaac catatgaatt ttctattgat tgtgacaaaa
     3301 taaacttatt ccgtggtgtc tttgcgtttc ttttatatgt tgccaccttt atgtatgtat
     3361 tttctacgtt tgctaacata ctgcgtaata aggagtctta atcatgccag ttcttttggc
     3421 tagcgccgcc ctataccttg tctgcctccc cgcgttgcgt cgcggtgcat ggagccgggc
     3481 cacctcgacc tgaatggaag ccggcggcac ctcgctaacg gattcaccac tccaagaatt
     3541 ggagccaatc aattcttgcg gagaactgtg aatgcgcaaa ccaacccttg gcagaacata
     3601 tccatcgcgt ccgccatctc cagcagccgc acgcggcgca tctcgggcag cgttgggtcc
     3661 tggccacggg tgcgcatgat cgtgctcctg tcgttgagga cccggctagg ctggcggggt
     3721 tgccttactg gttagcagaa tgaatcaccg atacgcgagc gaacgtgaag cgactgctgc
     3781 tgcaaaacgt ctgcgacctg agcaacaaca tgaatggtct tcggtttccg tgtttcgtaa
     3841 agtctggaaa cgcggaagtc agcgccctgc accattatgt tccggatctg catcgcagga
     3901 tgctgctggc taccctgtgg aacacctaca tctgtattaa cgaagcgctg gcattgaccc
     3961 tgagtgattt ttctctggtc ccgccgcatc cataccgcca gttgtttacc ctcacaacgt
     4021 tccagtaacc gggcatgttc atcatcagta acccgtatcg tgagcatcct ctctcgtttc
     4081 atcggtatca ttacccccat gaacagaaat cccccttaca cggaggcatc agtgaccaaa
     4141 caggaaaaaa ccgcccttaa catggcccgc tttatcagaa gccagacatt aacgcttctg
     4201 gagaaactca acgagctgga cgcggatgaa caggcagaca tctgtgaatc gcttcacgac
     4261 cacgctgatg agctttaccg caggatccgc gaaattgtaa gcgttaatat tttgttaaaa
     4321 ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa
     4381 atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac
     4441 aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag
     4501 ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt
     4561 aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg
     4621 gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca
     4681 agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag
     4741 ggcgcgtcgc ggatcctgcc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     4801 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     4861 tcagggcgcg tcagcgggtg ttggcgggtg tcggggcgca gccatgaccc agtcacgtag
     4921 cgatagcgga gtgtatactg gcttaactat gcggcatcag agcagattgt actgagagtg
     4981 caccatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc
     5041 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     5101 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     5161 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     5221 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     5281 tggcgaaacc cgacaggact ataaagatac cagggtttcc ccctggaagc tccctcgtgc
     5341 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     5401 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     5461 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     5521 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     5581 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     5641 ctaactacgg ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta
     5701 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     5761 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     5821 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     5881 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     5941 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     6001 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     6061 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     6121 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     6181 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     6241 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctgcag
     6301 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     6361 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     6421 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     6481 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     6541 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaacac
     6601 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6661 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     6721 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     6781 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     6841 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     6901 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     6961 aagtgccacc tgacgtctaa gaaaccatta ttatcatgac at
//
