LOCUS       sequence-191066-        6894 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            SH3D19-2/5.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6894)
  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 6894)
  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_SH3D19-2/5
FEATURES             Location/Qualifiers
     source          1..6894
                     /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      2544..2591
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4184..4639)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4321..4342)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4533..4552
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4681..4703)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4789..4929
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4844..4863
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4944..4961)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5115..5702)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5873..6664,6665..6733))
                     /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     6496..6515
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6734..6838)
                     /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 ccgcacgctg tcgtcctgca tgacttcccg gctgaacaag
     2401 tggatgacct gaatctgacc tctggcgaaa ttgtctatct gctggaaaaa attgataccg
     2461 actggtatcg tggcaactgc cgcaatcaga ttggtatctt tccggcgaac tacgtgaaag
     2521 ttattatcgc ggccgcataa taaaccgata caattaaagg ctccttttgg agcctttttt
     2581 ttggagattt tcaacgtgaa aaaattatta ttcgcaattc ctttagttgt tcctttctat
     2641 tctcactccg ctgaaactgt tgaaagttgt ttagcaaaac cccatacaga aaattcattt
     2701 actaacgtct ggaaagacga caaaacttta gatcgttacg ctaactatga gggctgtctg
     2761 tggaatgcta caggcgttgt agtttgtact ggtgacgaaa ctcagtgtct agctagagtg
     2821 gcggtggctc tggttccggt gattttgatt atgaaaagat ggcaaacgct aataaggggg
     2881 ctatgaccga aaatgccgat gaaaacgcgc tacagtctga cgctaaaggc aaacttgatt
     2941 ctgtcgctac tgattacggt gctgctatcg atggtttcat tggtgacgtt tccggccttg
     3001 ctaatggtaa tggtgctact ggtgattttg ctggctctaa ttcccaaatg gctcaagtcg
     3061 gtgacggtga taattcacct ttaatgaata atttccgtca atatttacct tccctccctc
     3121 aatcggttga atgtcgccct tttgtctttg gcgctggtaa accatatgaa ttttctattg
     3181 attgtgacaa aataaactta ttccgtggtg tctttgcgtt tcttttatat gttgccacct
     3241 ttatgtatgt attttctacg tttgctaaca tactgcgtaa taaggagtct taatcatgcc
     3301 agttcttttg gctagcgccg ccctatacct tgtctgcctc cccgcgttgc gtcgcggtgc
     3361 atggagccgg gccacctcga cctgaatgga agccggcggc acctcgctaa cggattcacc
     3421 actccaagaa ttggagccaa tcaattcttg cggagaactg tgaatgcgca aaccaaccct
     3481 tggcagaaca tatccatcgc gtccgccatc tccagcagcc gcacgcggcg catctcgggc
     3541 agcgttgggt cctggccacg ggtgcgcatg atcgtgctcc tgtcgttgag gacccggcta
     3601 ggctggcggg gttgccttac tggttagcag aatgaatcac cgatacgcga gcgaacgtga
     3661 agcgactgct gctgcaaaac gtctgcgacc tgagcaacaa catgaatggt cttcggtttc
     3721 cgtgtttcgt aaagtctgga aacgcggaag tcagcgccct gcaccattat gttccggatc
     3781 tgcatcgcag gatgctgctg gctaccctgt ggaacaccta catctgtatt aacgaagcgc
     3841 tggcattgac cctgagtgat ttttctctgg tcccgccgca tccataccgc cagttgttta
     3901 ccctcacaac gttccagtaa ccgggcatgt tcatcatcag taacccgtat cgtgagcatc
     3961 ctctctcgtt tcatcggtat cattaccccc atgaacagaa atccccctta cacggaggca
     4021 tcagtgacca aacaggaaaa aaccgccctt aacatggccc gctttatcag aagccagaca
     4081 ttaacgcttc tggagaaact caacgagctg gacgcggatg aacaggcaga catctgtgaa
     4141 tcgcttcacg accacgctga tgagctttac cgcaggatcc gcgaaattgt aagcgttaat
     4201 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc
     4261 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt
     4321 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa
     4381 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg
     4441 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga
     4501 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct
     4561 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat
     4621 gcgccgctac agggcgcgtc gcggatcctg cctcgcgcgt ttcggtgatg acggtgaaaa
     4681 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     4741 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac
     4801 ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt
     4861 gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     4921 cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     4981 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     5041 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     5101 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     5161 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accagggttt ccccctggaa
     5221 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     5281 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     5341 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     5401 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     5461 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     5521 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     5581 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     5641 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     5701 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     5761 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     5821 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     5881 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     5941 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     6001 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     6061 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     6121 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     6181 ccattgctgc aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     6241 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     6301 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     6361 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     6421 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     6481 cggcgtcaac acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     6541 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     6601 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     6661 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     6721 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     6781 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     6841 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acat
//
