LOCUS       sequence-192291-        6931 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            ITSN1-1/5-IS1/3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6931)
  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 6931)
  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_ITSN1-1/5-IS1/3
FEATURES             Location/Qualifiers
     source          1..6931
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        41..118
                     /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             119..1201
                     /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(138..157)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1444..1449,1450..1465,1466..1472)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1480..1496
                     /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     1485..1507
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1581..1598
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1614..2267
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2225..2247
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2274..2297
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2310..2327
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2571..2618
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4211..4666)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4348..4369)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4560..4579
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4708..4730)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4816..4954
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4871..4890
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4969..4986)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5140..5727)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5898..6689,6690..6758))
                     /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     6521..6540
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6759..6863)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     join(6931,1..18)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ataggcgtat cacgaggccc tttcgtcttc aagaattccc gacaccatcg aatggtgcaa
       61 aacctttcgc ggtatggcat gatagcgccc ggaagagagt caattcaggg tggtgaatgt
      121 gaaaccagta acgttatacg atgtcgcaga gtatgccggt gtctcttatc agaccgtttc
      181 ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc
      241 gatggcggag ctgaattaca ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc
      301 gttgctgatt ggcgttgcca cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc
      361 ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg gtggtgtcga tggtagaacg
      421 aagcggcgtc gaagcctgta aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg
      481 gctgatcatt aactatccgc tggatgacca ggatgccatt gctgtggaag ctgcctgcac
      541 taatgttccg gcgttatttc ttgatgtctc tgaccagaca cccatcaaca gtattatttt
      601 ctcccatgaa gacggtacgc gactgggcgt ggagcatctg gtcgcattgg gtcaccagca
      661 aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg cgtctgcgtc tggctggctg
      721 gcataaatat ctcactcgca atcaaattca gccgatagcg gaacgggaag gcgactggag
      781 tgccatgtcc ggttttcaac aaaccatgca aatgctgaat gagggcatcg ttcccactgc
      841 gatgctggtt gccaacgatc agatggcgct gggcgcaatg cgcgccatta ccgagtccgg
      901 gctgcgcgtt ggtgcggata tctcggtagt gggatacgac gataccgaag acagctcatg
      961 ttatatcccg ccgttaacca ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt
     1021 ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt
     1081 ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg caaaccgcct ctccccgcgc
     1141 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     1201 agcgcaacgc aattaatgta agttagctca ctcattaggc acaattctca tgtttgacag
     1261 cttatcatcg actgcacggt gcaccaatgc ttctggcgtc aggcagccat cggaagctgt
     1321 ggtatggctg tgcaggtcgt aaatcactgc ataattcgtg tcgctcaagg cgcactcccg
     1381 ttctggataa tgttttttgc gccgacatca taacggttct ggcaaatatt ctgaaatgag
     1441 ctgttgacaa ttaatcatcg gctcgtataa tgtgtggaat tgtgagcgga taacaatttc
     1501 acacaggaaa cagccagtcc gtttaggtgt tttcacgagc acttcaccaa caaggaccat
     1561 agattatgaa aatcgaagaa caccatcacc atcaccattc cagcggtaag cttatgtccc
     1621 ctatactagg ttattggaaa attaagggcc ttgtgcaacc cactcgactt cttttggaat
     1681 atcttgaaga aaaatatgaa gagcatttgt atgagcgcga tgaaggtgat aaatggcgaa
     1741 acaaaaagtt tgaattgggt ttggagtttc ccaatcttcc ttattatatt gatggtgatg
     1801 ttaaattaac acagtctatg gccatcatac gttatatagc tgacaagcac aacatgttgg
     1861 gtggttgtcc aaaagagcgt gcagagattt caatgcttga aggagcggtt ttggatatta
     1921 gatacggtgt ttcgagaatt gcatatagta aagactttga aactctcaaa gttgattttc
     1981 ttagcaagct acctgaaatg ctgaaaatgt tcgaagatcg tttatgtcat aaaacatatt
     2041 taaatggtga tcatgtaacc catcctgact tcatgttgta tgacgctctt gatgttgttt
     2101 tatacatgga cccaatgtgc ctggatgcgt tcccaaaatt agtttgtttt aaaaaacgta
     2161 ttgaagctat cccacaaatt gataagtact tgaaatccag caagtatata gcatggcctt
     2221 tgcagggctg gcaagccacg tttggtggtg gcgaccatcc tccaaaatcg gatctggaag
     2281 ttctgttcca ggggcccctg tccagcggtc tggttccgcg tggttccggt accgcggccc
     2341 agccggccat ggcccagggc gcgctggttt actatcgtgc cctgtaccca tttgaatccc
     2401 gttcccatga tgaaatcacc attcagccgg gtgatattgt catggttaaa ggtgaatggg
     2461 ttgacgaatc ccaaactggt gaaccaggtt ggctgggtgg tgaactgaaa ggtaaaaccg
     2521 gttggtttcc agcaaactat gccgaaaaaa ttccggcggc cgcataataa accgatacaa
     2581 ttaaaggctc cttttggagc cttttttttg gagattttca acgtgaaaaa attattattc
     2641 gcaattcctt tagttgttcc tttctattct cactccgctg aaactgttga aagttgttta
     2701 gcaaaacccc atacagaaaa ttcatttact aacgtctgga aagacgacaa aactttagat
     2761 cgttacgcta actatgaggg ctgtctgtgg aatgctacag gcgttgtagt ttgtactggt
     2821 gacgaaactc agtgtctagc tagagtggcg gtggctctgg ttccggtgat tttgattatg
     2881 aaaagatggc aaacgctaat aagggggcta tgaccgaaaa tgccgatgaa aacgcgctac
     2941 agtctgacgc taaaggcaaa cttgattctg tcgctactga ttacggtgct gctatcgatg
     3001 gtttcattgg tgacgtttcc ggccttgcta atggtaatgg tgctactggt gattttgctg
     3061 gctctaattc ccaaatggct caagtcggtg acggtgataa ttcaccttta atgaataatt
     3121 tccgtcaata tttaccttcc ctccctcaat cggttgaatg tcgccctttt gtctttggcg
     3181 ctggtaaacc atatgaattt tctattgatt gtgacaaaat aaacttattc cgtggtgtct
     3241 ttgcgtttct tttatatgtt gccaccttta tgtatgtatt ttctacgttt gctaacatac
     3301 tgcgtaataa ggagtcttaa tcatgccagt tcttttggct agcgccgccc tataccttgt
     3361 ctgcctcccc gcgttgcgtc gcggtgcatg gagccgggcc acctcgacct gaatggaagc
     3421 cggcggcacc tcgctaacgg attcaccact ccaagaattg gagccaatca attcttgcgg
     3481 agaactgtga atgcgcaaac caacccttgg cagaacatat ccatcgcgtc cgccatctcc
     3541 agcagccgca cgcggcgcat ctcgggcagc gttgggtcct ggccacgggt gcgcatgatc
     3601 gtgctcctgt cgttgaggac ccggctaggc tggcggggtt gccttactgg ttagcagaat
     3661 gaatcaccga tacgcgagcg aacgtgaagc gactgctgct gcaaaacgtc tgcgacctga
     3721 gcaacaacat gaatggtctt cggtttccgt gtttcgtaaa gtctggaaac gcggaagtca
     3781 gcgccctgca ccattatgtt ccggatctgc atcgcaggat gctgctggct accctgtgga
     3841 acacctacat ctgtattaac gaagcgctgg cattgaccct gagtgatttt tctctggtcc
     3901 cgccgcatcc ataccgccag ttgtttaccc tcacaacgtt ccagtaaccg ggcatgttca
     3961 tcatcagtaa cccgtatcgt gagcatcctc tctcgtttca tcggtatcat tacccccatg
     4021 aacagaaatc ccccttacac ggaggcatca gtgaccaaac aggaaaaaac cgcccttaac
     4081 atggcccgct ttatcagaag ccagacatta acgcttctgg agaaactcaa cgagctggac
     4141 gcggatgaac aggcagacat ctgtgaatcg cttcacgacc acgctgatga gctttaccgc
     4201 aggatccgcg aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
     4261 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
     4321 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
     4381 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
     4441 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
     4501 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
     4561 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
     4621 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcgcg gatcctgcct
     4681 cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac
     4741 agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt
     4801 tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag tgtatactgg
     4861 cttaactatg cggcatcaga gcagattgta ctgagagtgc actatgcggt gtgaaatacc
     4921 gcacagatgc gtaaggagaa aataccgcat caggcgctct tccgcttcct cgctcactga
     4981 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     5041 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     5101 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     5161 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     5221 aagataccag ggtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     5281 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     5341 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     5401 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     5461 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     5521 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg
     5581 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     5641 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     5701 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     5761 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     5821 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     5881 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     5941 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     6001 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     6061 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     6121 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     6181 gttaatagtt tgcgcaacgt tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg
     6241 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     6301 atattgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     6361 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     6421 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     6481 atgcggcgac cgagttgctc ttgcccggcg tcaacacggg ataataccgc gccacatagc
     6541 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     6601 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     6661 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     6721 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     6781 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     6841 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa
     6901 accattatta tcatgacatt aacctataaa a
//
