LOCUS       sequence-192248-        6948 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            OTOR-2/2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6948)
  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 6948)
  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_OTOR-2/2
FEATURES             Location/Qualifiers
     source          1..6948
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        37..114
                     /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             115..1197
                     /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(134..153)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1440..1445,1446..1461,1462..1468)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1476..1492
                     /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     1481..1503
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1577..1594
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1610..2263
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2221..2243
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2270..2293
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2306..2323
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2582..2629
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4222..4677)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4359..4380)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4571..4590
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4719..4741)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4827..4967
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4882..4901
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4982..4999)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5153..5740)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5911..6702,6703..6771))
                     /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     6534..6553
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6772..6876)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     join(6944..6948,1..14)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gcgtatcacg aggccctttc gtcttcaaga attcccgaca ccatcgaatg gtgcaaaacc
       61 tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt gaatgtgaaa
      121 ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac cgtttcccgc
      181 gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga agcggcgatg
      241 gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa acagtcgttg
      301 ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat tgtcgcggcg
      361 attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt agaacgaagc
      421 ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt cagtgggctg
      481 atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc ctgcactaat
      541 gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat tattttctcc
      601 catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca ccagcaaatc
      661 gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc tggctggcat
      721 aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga ctggagtgcc
      781 atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc cactgcgatg
      841 ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga gtccgggctg
      901 cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag ctcatgttat
      961 atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac cagcgtggac
     1021 cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt gcccgtctca
     1081 ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
     1141 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg
     1201 caacgcaatt aatgtaagtt agctcactca ttaggcacaa ttctcatgtt tgacagctta
     1261 tcatcgactg cacggtgcac caatgcttct ggcgtcaggc agccatcgga agctgtggta
     1321 tggctgtgca ggtcgtaaat cactgcataa ttcgtgtcgc tcaaggcgca ctcccgttct
     1381 ggataatgtt ttttgcgccg acatcataac ggttctggca aatattctga aatgagctgt
     1441 tgacaattaa tcatcggctc gtataatgtg tggaattgtg agcggataac aatttcacac
     1501 aggaaacagc cagtccgttt aggtgttttc acgagcactt caccaacaag gaccatagat
     1561 tatgaaaatc gaagaacacc atcaccatca ccattccagc ggtaagctta tgtcccctat
     1621 actaggttat tggaaaatta agggccttgt gcaacccact cgacttcttt tggaatatct
     1681 tgaagaaaaa tatgaagagc atttgtatga gcgcgatgaa ggtgataaat ggcgaaacaa
     1741 aaagtttgaa ttgggtttgg agtttcccaa tcttccttat tatattgatg gtgatgttaa
     1801 attaacacag tctatggcca tcatacgtta tatagctgac aagcacaaca tgttgggtgg
     1861 ttgtccaaaa gagcgtgcag agatttcaat gcttgaagga gcggttttgg atattagata
     1921 cggtgtttcg agaattgcat atagtaaaga ctttgaaact ctcaaagttg attttcttag
     1981 caagctacct gaaatgctga aaatgttcga agatcgttta tgtcataaaa catatttaaa
     2041 tggtgatcat gtaacccatc ctgacttcat gttgtatgac gctcttgatg ttgttttata
     2101 catggaccca atgtgcctgg atgcgttccc aaaattagtt tgttttaaaa aacgtattga
     2161 agctatccca caaattgata agtacttgaa atccagcaag tatatagcat ggcctttgca
     2221 gggctggcaa gccacgtttg gtggtggcga ccatcctcca aaatcggatc tggaagttct
     2281 gttccagggg cccctgtcca gcggtctggt tccgcgtggt tccggtaccg cggcccagcc
     2341 ggccatggcc cagggcgcgc tgagtctggc atccgcccag gaagattaca atgccccgga
     2401 ttgtcgtttc attaacgtca aaaaaggtca gcagatttac gtttactcca aactggtcaa
     2461 agaaaacggt gccggtgaat tttgggcagg ttccgtttat ggtgatggtc aggatgaaat
     2521 gggtgtcgtt ggttactttc cacgtaacct ggttaaagaa cagcgtgcgg ccgcataata
     2581 aaccgataca attaaaggct ccttttggag cctttttttt ggagattttc aacgtgaaaa
     2641 aattattatt cgcaattcct ttagttgttc ctttctattc tcactccgct gaaactgttg
     2701 aaagttgttt agcaaaaccc catacagaaa attcatttac taacgtctgg aaagacgaca
     2761 aaactttaga tcgttacgct aactatgagg gctgtctgtg gaatgctaca ggcgttgtag
     2821 tttgtactgg tgacgaaact cagtgtctag ctagagtggc ggtggctctg gttccggtga
     2881 ttttgattat gaaaagatgg caaacgctaa taagggggct atgaccgaaa atgccgatga
     2941 aaacgcgcta cagtctgacg ctaaaggcaa acttgattct gtcgctactg attacggtgc
     3001 tgctatcgat ggtttcattg gtgacgtttc cggccttgct aatggtaatg gtgctactgg
     3061 tgattttgct ggctctaatt cccaaatggc tcaagtcggt gacggtgata attcaccttt
     3121 aatgaataat ttccgtcaat atttaccttc cctccctcaa tcggttgaat gtcgcccttt
     3181 tgtctttggc gctggtaaac catatgaatt ttctattgat tgtgacaaaa taaacttatt
     3241 ccgtggtgtc tttgcgtttc ttttatatgt tgccaccttt atgtatgtat tttctacgtt
     3301 tgctaacata ctgcgtaata aggagtctta atcatgccag ttcttttggc tagcgccgcc
     3361 ctataccttg tctgcctccc cgcgttgcgt cgcggtgcat ggagccgggc cacctcgacc
     3421 tgaatggaag ccggcggcac ctcgctaacg gattcaccac tccaagaatt ggagccaatc
     3481 aattcttgcg gagaactgtg aatgcgcaaa ccaacccttg gcagaacata tccatcgcgt
     3541 ccgccatctc cagcagccgc acgcggcgca tctcgggcag cgttgggtcc tggccacggg
     3601 tgcgcatgat cgtgctcctg tcgttgagga cccggctagg ctggcggggt tgccttactg
     3661 gttagcagaa tgaatcaccg atacgcgagc gaacgtgaag cgactgctgc tgcaaaacgt
     3721 ctgcgacctg agcaacaaca tgaatggtct tcggtttccg tgtttcgtaa agtctggaaa
     3781 cgcggaagtc agcgccctgc accattatgt tccggatctg catcgcagga tgctgctggc
     3841 taccctgtgg aacacctaca tctgtattaa cgaagcgctg gcattgaccc tgagtgattt
     3901 ttctctggtc ccgccgcatc cataccgcca gttgtttacc ctcacaacgt tccagtaacc
     3961 gggcatgttc atcatcagta acccgtatcg tgagcatcct ctctcgtttc atcggtatca
     4021 ttacccccat gaacagaaat cccccttaca cggaggcatc agtgaccaaa caggaaaaaa
     4081 ccgcccttaa catggcccgc tttatcagaa gccagacatt aacgcttctg gagaaactca
     4141 acgagctgga cgcggatgaa caggcagaca tctgtgaatc gcttcacgac cacgctgatg
     4201 agctttaccg caggatccgc gaaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa
     4261 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
     4321 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
     4381 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
     4441 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
     4501 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
     4561 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
     4621 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcgc
     4681 ggatcctgcc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
     4741 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
     4801 tcagcgggtg ttggcgggtg tcggggcgca gccatgaccc agtcacgtag cgatagcgga
     4861 gtgtatactg gcttaactat gcggcatcag agcagattgt actgagagtg caccatatgc
     4921 ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc tcttccgctt
     4981 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     5041 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     5101 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     5161 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     5221 cgacaggact ataaagatac cagggtttcc ccctggaagc tccctcgtgc gctctcctgt
     5281 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     5341 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     5401 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     5461 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     5521 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     5581 ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     5641 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     5701 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     5761 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     5821 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     5881 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     5941 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     6001 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     6061 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     6121 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     6181 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctgcag gcatcgtggt
     6241 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     6301 tacatgatcc cccatattgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     6361 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     6421 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     6481 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaacac gggataatac
     6541 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     6601 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     6661 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     6721 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     6781 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     6841 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     6901 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaatag
//
