LOCUS       sequence-191061-        6927 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            MPP3-1/1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6927)
  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 6927)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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_MPP3-1/1
FEATURES             Location/Qualifiers
     source          1..6927
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     5..23
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        46..123
                     /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             124..1206
                     /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(143..162)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1449..1454,1455..1470,1471..1477)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1485..1501
                     /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     1490..1512
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1586..1603
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1619..2272
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2230..2252
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2279..2302
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2315..2332
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2570..2617
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4210..4665)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4347..4368)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4559..4578
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4707..4729)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4815..4955
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4870..4889
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4970..4987)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5141..5728)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5899..6690,6691..6759))
                     /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     6522..6541
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6760..6864)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 taaaaatagg cgtatcacga ggccctttcg tcttcaagaa ttcccgacac catcgaatgg
       61 tgcaaaacct ttcgcggtat ggcatgatag cgcccggaag agagtcaatt cagggtggtg
      121 aatgtgaaac cagtaacgtt atacgatgtc gcagagtatg ccggtgtctc ttatcagacc
      181 gtttcccgcg tggtgaacca ggccagccac gtttctgcga aaacgcggga aaaagtggaa
      241 gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg cacaacaact ggcgggcaaa
      301 cagtcgttgc tgattggcgt tgccacctcc agtctggccc tgcacgcgcc gtcgcaaatt
      361 gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca gcgtggtggt gtcgatggta
      421 gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca atcttctcgc gcaacgcgtc
      481 agtgggctga tcattaacta tccgctggat gaccaggatg ccattgctgt ggaagctgcc
      541 tgcactaatg ttccggcgtt atttcttgat gtctctgacc agacacccat caacagtatt
      601 attttctccc atgaagacgg tacgcgactg ggcgtggagc atctggtcgc attgggtcac
      661 cagcaaatcg cgctgttagc gggcccatta agttctgtct cggcgcgtct gcgtctggct
      721 ggctggcata aatatctcac tcgcaatcaa attcagccga tagcggaacg ggaaggcgac
      781 tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg catcgttccc
      841 actgcgatgc tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc cattaccgag
      901 tccgggctgc gcgttggtgc ggatatctcg gtagtgggat acgacgatac cgaagacagc
      961 tcatgttata tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct ggggcaaacc
     1021 agcgtggacc gcttgctgca actctctcag ggccaggcgg tgaagggcaa tcagctgttg
     1081 cccgtctcac tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac cgcctctccc
     1141 cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg
     1201 cagtgagcgc aacgcaatta atgtaagtta gctcactcat taggcacaat tctcatgttt
     1261 gacagcttat catcgactgc acggtgcacc aatgcttctg gcgtcaggca gccatcggaa
     1321 gctgtggtat ggctgtgcag gtcgtaaatc actgcataat tcgtgtcgct caaggcgcac
     1381 tcccgttctg gataatgttt tttgcgccga catcataacg gttctggcaa atattctgaa
     1441 atgagctgtt gacaattaat catcggctcg tataatgtgt ggaattgtga gcggataaca
     1501 atttcacaca ggaaacagcc agtccgttta ggtgttttca cgagcacttc accaacaagg
     1561 accatagatt atgaaaatcg aagaacacca tcaccatcac cattccagcg gtaagcttat
     1621 gtcccctata ctaggttatt ggaaaattaa gggccttgtg caacccactc gacttctttt
     1681 ggaatatctt gaagaaaaat atgaagagca tttgtatgag cgcgatgaag gtgataaatg
     1741 gcgaaacaaa aagtttgaat tgggtttgga gtttcccaat cttccttatt atattgatgg
     1801 tgatgttaaa ttaacacagt ctatggccat catacgttat atagctgaca agcacaacat
     1861 gttgggtggt tgtccaaaag agcgtgcaga gatttcaatg cttgaaggag cggttttgga
     1921 tattagatac ggtgtttcga gaattgcata tagtaaagac tttgaaactc tcaaagttga
     1981 ttttcttagc aagctacctg aaatgctgaa aatgttcgaa gatcgtttat gtcataaaac
     2041 atatttaaat ggtgatcatg taacccatcc tgacttcatg ttgtatgacg ctcttgatgt
     2101 tgttttatac atggacccaa tgtgcctgga tgcgttccca aaattagttt gttttaaaaa
     2161 acgtattgaa gctatcccac aaattgataa gtacttgaaa tccagcaagt atatagcatg
     2221 gcctttgcag ggctggcaag ccacgtttgg tggtggcgac catcctccaa aatcggatct
     2281 ggaagttctg ttccaggggc ccctgtccag cggtctggtt ccgcgtggtt ccggtaccgc
     2341 ggcccagccg gccgtgttta tgcgtgccct gttccattac aatccgcgtg aagatcgtgc
     2401 cattccgtgt caagaagcag gtctgccgtt ccagcgtcgt caggtcctgg aagtggttag
     2461 ccaagatgac ccgacctggt ggcaggcgaa acgtgttggc gatacgaacc tgcgcgccgg
     2521 cctgattccg tctaaaggtt ttcaagaacg tcgcgcggcc gcataataaa ccgatacaat
     2581 taaaggctcc ttttggagcc ttttttttgg agattttcaa cgtgaaaaaa ttattattcg
     2641 caattccttt agttgttcct ttctattctc actccgctga aactgttgaa agttgtttag
     2701 caaaacccca tacagaaaat tcatttacta acgtctggaa agacgacaaa actttagatc
     2761 gttacgctaa ctatgagggc tgtctgtgga atgctacagg cgttgtagtt tgtactggtg
     2821 acgaaactca gtgtctagct agagtggcgg tggctctggt tccggtgatt ttgattatga
     2881 aaagatggca aacgctaata agggggctat gaccgaaaat gccgatgaaa acgcgctaca
     2941 gtctgacgct aaaggcaaac ttgattctgt cgctactgat tacggtgctg ctatcgatgg
     3001 tttcattggt gacgtttccg gccttgctaa tggtaatggt gctactggtg attttgctgg
     3061 ctctaattcc caaatggctc aagtcggtga cggtgataat tcacctttaa tgaataattt
     3121 ccgtcaatat ttaccttccc tccctcaatc ggttgaatgt cgcccttttg tctttggcgc
     3181 tggtaaacca tatgaatttt ctattgattg tgacaaaata aacttattcc gtggtgtctt
     3241 tgcgtttctt ttatatgttg ccacctttat gtatgtattt tctacgtttg ctaacatact
     3301 gcgtaataag gagtcttaat catgccagtt cttttggcta gcgccgccct ataccttgtc
     3361 tgcctccccg cgttgcgtcg cggtgcatgg agccgggcca cctcgacctg aatggaagcc
     3421 ggcggcacct cgctaacgga ttcaccactc caagaattgg agccaatcaa ttcttgcgga
     3481 gaactgtgaa tgcgcaaacc aacccttggc agaacatatc catcgcgtcc gccatctcca
     3541 gcagccgcac gcggcgcatc tcgggcagcg ttgggtcctg gccacgggtg cgcatgatcg
     3601 tgctcctgtc gttgaggacc cggctaggct ggcggggttg ccttactggt tagcagaatg
     3661 aatcaccgat acgcgagcga acgtgaagcg actgctgctg caaaacgtct gcgacctgag
     3721 caacaacatg aatggtcttc ggtttccgtg tttcgtaaag tctggaaacg cggaagtcag
     3781 cgccctgcac cattatgttc cggatctgca tcgcaggatg ctgctggcta ccctgtggaa
     3841 cacctacatc tgtattaacg aagcgctggc attgaccctg agtgattttt ctctggtccc
     3901 gccgcatcca taccgccagt tgtttaccct cacaacgttc cagtaaccgg gcatgttcat
     3961 catcagtaac ccgtatcgtg agcatcctct ctcgtttcat cggtatcatt acccccatga
     4021 acagaaatcc cccttacacg gaggcatcag tgaccaaaca ggaaaaaacc gcccttaaca
     4081 tggcccgctt tatcagaagc cagacattaa cgcttctgga gaaactcaac gagctggacg
     4141 cggatgaaca ggcagacatc tgtgaatcgc ttcacgacca cgctgatgag ctttaccgca
     4201 ggatccgcga aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     4261 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     4321 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     4381 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     4441 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     4501 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     4561 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     4621 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcgcgg atcctgcctc
     4681 gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca
     4741 gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt
     4801 ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc
     4861 ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac
     4921 cgcacagatg cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc tcgctcactg
     4981 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     5041 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     5101 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     5161 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     5221 aaagatacca gggtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     5281 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     5341 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     5401 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     5461 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     5521 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     5581 gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     5641 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     5701 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     5761 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     5821 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     5881 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     5941 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     6001 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     6061 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     6121 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     6181 agttaatagt ttgcgcaacg ttgttgccat tgctgcaggc atcgtggtgt cacgctcgtc
     6241 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     6301 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     6361 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     6421 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     6481 tatgcggcga ccgagttgct cttgcccggc gtcaacacgg gataataccg cgccacatag
     6541 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     6601 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     6661 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     6721 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     6781 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     6841 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga
     6901 aaccattatt atcatgacat taaccta
//
