LOCUS       sequence-191146-        6975 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            MPP4-1/1-IS2/3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6975)
  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 6975)
  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_MPP4-1/1-IS2/3
FEATURES             Location/Qualifiers
     source          1..6975
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     33..51
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        74..151
                     /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             152..1234
                     /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(171..190)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1477..1482,1483..1498,1499..1505)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1513..1529
                     /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     1518..1540
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1614..1631
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1647..2300
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2258..2280
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2307..2330
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2343..2360
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2646..2693
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4286..4741)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4423..4444)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4635..4654
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4783..4805)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4891..5031
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4946..4965
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5046..5063)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5217..5804)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5975..6766,6767..6835))
                     /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     6598..6617
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6836..6940)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc
       61 ttcaagaatt cccgacacca tcgaatggtg caaaaccttt cgcggtatgg catgatagcg
      121 cccggaagag agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc
      181 agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt
      241 ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa
      301 ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag
      361 tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact
      421 gggtgccagc gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc
      481 ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga
      541 ccaggatgcc attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt
      601 ctctgaccag acacccatca acagtattat tttctcccat gaagacggta cgcgactggg
      661 cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag
      721 ttctgtctcg gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat
      781 tcagccgata gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat
      841 gcaaatgctg aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc
      901 gctgggcgca atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt
      961 agtgggatac gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa
     1021 acaggatttt cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg
     1081 ccaggcggtg aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct
     1141 ggcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
     1201 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc
     1261 tcactcatta ggcacaattc tcatgtttga cagcttatca tcgactgcac ggtgcaccaa
     1321 tgcttctggc gtcaggcagc catcggaagc tgtggtatgg ctgtgcaggt cgtaaatcac
     1381 tgcataattc gtgtcgctca aggcgcactc ccgttctgga taatgttttt tgcgccgaca
     1441 tcataacggt tctggcaaat attctgaaat gagctgttga caattaatca tcggctcgta
     1501 taatgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagccag tccgtttagg
     1561 tgttttcacg agcacttcac caacaaggac catagattat gaaaatcgaa gaacaccatc
     1621 accatcacca ttccagcggt aagcttatgt cccctatact aggttattgg aaaattaagg
     1681 gccttgtgca acccactcga cttcttttgg aatatcttga agaaaaatat gaagagcatt
     1741 tgtatgagcg cgatgaaggt gataaatggc gaaacaaaaa gtttgaattg ggtttggagt
     1801 ttcccaatct tccttattat attgatggtg atgttaaatt aacacagtct atggccatca
     1861 tacgttatat agctgacaag cacaacatgt tgggtggttg tccaaaagag cgtgcagaga
     1921 tttcaatgct tgaaggagcg gttttggata ttagatacgg tgtttcgaga attgcatata
     1981 gtaaagactt tgaaactctc aaagttgatt ttcttagcaa gctacctgaa atgctgaaaa
     2041 tgttcgaaga tcgtttatgt cataaaacat atttaaatgg tgatcatgta acccatcctg
     2101 acttcatgtt gtatgacgct cttgatgttg ttttatacat ggacccaatg tgcctggatg
     2161 cgttcccaaa attagtttgt tttaaaaaac gtattgaagc tatcccacaa attgataagt
     2221 acttgaaatc cagcaagtat atagcatggc ctttgcaggg ctggcaagcc acgtttggtg
     2281 gtggcgacca tcctccaaaa tcggatctgg aagttctgtt ccaggggccc ctgtccagcg
     2341 gtctggttcc gcgtggttcc ggtaccgcgg cccagccggc cgacccgccg gttaactcgc
     2401 aacaaatggt ttacgttcgt gctatgaccg aatattggcc gcaagaagac ccggacatcc
     2461 cgtgtatgga tgcgggcctg ccgtttcaga aaggtgacat tctgcagatc gtcgatcaaa
     2521 acgacgcgct gtggtggcaa gcccgtaaaa ttagtgatcc ggcaacctgc gcaggtctgg
     2581 tgccgagcaa tcatctgctg aaacgctggt ctttcgcact ggttgctcag gcggccgcat
     2641 aataaaccga tacaattaaa ggctcctttt ggagcctttt ttttggagat tttcaacgtg
     2701 aaaaaattat tattcgcaat tcctttagtt gttcctttct attctcactc cgctgaaact
     2761 gttgaaagtt gtttagcaaa accccataca gaaaattcat ttactaacgt ctggaaagac
     2821 gacaaaactt tagatcgtta cgctaactat gagggctgtc tgtggaatgc tacaggcgtt
     2881 gtagtttgta ctggtgacga aactcagtgt ctagctagag tggcggtggc tctggttccg
     2941 gtgattttga ttatgaaaag atggcaaacg ctaataaggg ggctatgacc gaaaatgccg
     3001 atgaaaacgc gctacagtct gacgctaaag gcaaacttga ttctgtcgct actgattacg
     3061 gtgctgctat cgatggtttc attggtgacg tttccggcct tgctaatggt aatggtgcta
     3121 ctggtgattt tgctggctct aattcccaaa tggctcaagt cggtgacggt gataattcac
     3181 ctttaatgaa taatttccgt caatatttac cttccctccc tcaatcggtt gaatgtcgcc
     3241 cttttgtctt tggcgctggt aaaccatatg aattttctat tgattgtgac aaaataaact
     3301 tattccgtgg tgtctttgcg tttcttttat atgttgccac ctttatgtat gtattttcta
     3361 cgtttgctaa catactgcgt aataaggagt cttaatcatg ccagttcttt tggctagcgc
     3421 cgccctatac cttgtctgcc tccccgcgtt gcgtcgcggt gcatggagcc gggccacctc
     3481 gacctgaatg gaagccggcg gcacctcgct aacggattca ccactccaag aattggagcc
     3541 aatcaattct tgcggagaac tgtgaatgcg caaaccaacc cttggcagaa catatccatc
     3601 gcgtccgcca tctccagcag ccgcacgcgg cgcatctcgg gcagcgttgg gtcctggcca
     3661 cgggtgcgca tgatcgtgct cctgtcgttg aggacccggc taggctggcg gggttgcctt
     3721 actggttagc agaatgaatc accgatacgc gagcgaacgt gaagcgactg ctgctgcaaa
     3781 acgtctgcga cctgagcaac aacatgaatg gtcttcggtt tccgtgtttc gtaaagtctg
     3841 gaaacgcgga agtcagcgcc ctgcaccatt atgttccgga tctgcatcgc aggatgctgc
     3901 tggctaccct gtggaacacc tacatctgta ttaacgaagc gctggcattg accctgagtg
     3961 atttttctct ggtcccgccg catccatacc gccagttgtt taccctcaca acgttccagt
     4021 aaccgggcat gttcatcatc agtaacccgt atcgtgagca tcctctctcg tttcatcggt
     4081 atcattaccc ccatgaacag aaatccccct tacacggagg catcagtgac caaacaggaa
     4141 aaaaccgccc ttaacatggc ccgctttatc agaagccaga cattaacgct tctggagaaa
     4201 ctcaacgagc tggacgcgga tgaacaggca gacatctgtg aatcgcttca cgaccacgct
     4261 gatgagcttt accgcaggat ccgcgaaatt gtaagcgtta atattttgtt aaaattcgcg
     4321 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     4381 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     4441 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     4501 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     4561 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     4621 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     4681 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     4741 tcgcggatcc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
     4801 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     4861 cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag
     4921 cggagtgtat actggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     4981 atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag gcgctcttcc
     5041 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     5101 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     5161 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     5221 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     5281 aacccgacag gactataaag ataccagggt ttccccctgg aagctccctc gtgcgctctc
     5341 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     5401 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5461 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5521 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5581 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5641 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5701 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5761 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5821 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5881 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5941 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     6001 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     6061 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     6121 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     6181 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6241 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct gcaggcatcg
     6301 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6361 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6421 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6481 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6541 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca acacgggata
     6601 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6661 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     6721 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     6781 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     6841 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6901 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     6961 cacctgacgt ctaag
//
