LOCUS       sequence-191804-        7050 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            OTOR-1/2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7050)
  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 7050)
  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-1/2
FEATURES             Location/Qualifiers
     source          1..7050
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..19
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        42..119
                     /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             120..1202
                     /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(139..158)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1445..1450,1451..1466,1467..1473)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1481..1497
                     /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     1486..1508
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1582..1599
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1615..2268
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2226..2248
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2275..2298
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2311..2328
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2689..2736
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4329..4784)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4466..4487)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4678..4697
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4826..4848)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4934..5074
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4989..5008
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5089..5106)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5260..5847)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6018..6809,6810..6878))
                     /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     6641..6660
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6879..6983)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aataggcgta tcacgaggcc ctttcgtctt caagaattcc cgacaccatc gaatggtgca
       61 aaacctttcg cggtatggca tgatagcgcc cggaagagag tcaattcagg gtggtgaatg
      121 tgaaaccagt aacgttatac gatgtcgcag agtatgccgg tgtctcttat cagaccgttt
      181 cccgcgtggt gaaccaggcc agccacgttt ctgcgaaaac gcgggaaaaa gtggaagcgg
      241 cgatggcgga gctgaattac attcccaacc gcgtggcaca acaactggcg ggcaaacagt
      301 cgttgctgat tggcgttgcc acctccagtc tggccctgca cgcgccgtcg caaattgtcg
      361 cggcgattaa atctcgcgcc gatcaactgg gtgccagcgt ggtggtgtcg atggtagaac
      421 gaagcggcgt cgaagcctgt aaagcggcgg tgcacaatct tctcgcgcaa cgcgtcagtg
      481 ggctgatcat taactatccg ctggatgacc aggatgccat tgctgtggaa gctgcctgca
      541 ctaatgttcc ggcgttattt cttgatgtct ctgaccagac acccatcaac agtattattt
      601 tctcccatga agacggtacg cgactgggcg tggagcatct ggtcgcattg ggtcaccagc
      661 aaatcgcgct gttagcgggc ccattaagtt ctgtctcggc gcgtctgcgt ctggctggct
      721 ggcataaata tctcactcgc aatcaaattc agccgatagc ggaacgggaa ggcgactgga
      781 gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa tgagggcatc gttcccactg
      841 cgatgctggt tgccaacgat cagatggcgc tgggcgcaat gcgcgccatt accgagtccg
      901 ggctgcgcgt tggtgcggat atctcggtag tgggatacga cgataccgaa gacagctcat
      961 gttatatccc gccgttaacc accatcaaac aggattttcg cctgctgggg caaaccagcg
     1021 tggaccgctt gctgcaactc tctcagggcc aggcggtgaa gggcaatcag ctgttgcccg
     1081 tctcactggt gaaaagaaaa accaccctgg cgcccaatac gcaaaccgcc tctccccgcg
     1141 cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt
     1201 gagcgcaacg caattaatgt aagttagctc actcattagg cacaattctc atgtttgaca
     1261 gcttatcatc gactgcacgg tgcaccaatg cttctggcgt caggcagcca tcggaagctg
     1321 tggtatggct gtgcaggtcg taaatcactg cataattcgt gtcgctcaag gcgcactccc
     1381 gttctggata atgttttttg cgccgacatc ataacggttc tggcaaatat tctgaaatga
     1441 gctgttgaca attaatcatc ggctcgtata atgtgtggaa ttgtgagcgg ataacaattt
     1501 cacacaggaa acagccagtc cgtttaggtg ttttcacgag cacttcacca acaaggacca
     1561 tagattatga aaatcgaaga acaccatcac catcaccatt ccagcggtaa gcttatgtcc
     1621 cctatactag gttattggaa aattaagggc cttgtgcaac ccactcgact tcttttggaa
     1681 tatcttgaag aaaaatatga agagcatttg tatgagcgcg atgaaggtga taaatggcga
     1741 aacaaaaagt ttgaattggg tttggagttt cccaatcttc cttattatat tgatggtgat
     1801 gttaaattaa cacagtctat ggccatcata cgttatatag ctgacaagca caacatgttg
     1861 ggtggttgtc caaaagagcg tgcagagatt tcaatgcttg aaggagcggt tttggatatt
     1921 agatacggtg tttcgagaat tgcatatagt aaagactttg aaactctcaa agttgatttt
     1981 cttagcaagc tacctgaaat gctgaaaatg ttcgaagatc gtttatgtca taaaacatat
     2041 ttaaatggtg atcatgtaac ccatcctgac ttcatgttgt atgacgctct tgatgttgtt
     2101 ttatacatgg acccaatgtg cctggatgcg ttcccaaaat tagtttgttt taaaaaacgt
     2161 attgaagcta tcccacaaat tgataagtac ttgaaatcca gcaagtatat agcatggcct
     2221 ttgcagggct ggcaagccac gtttggtggt ggcgaccatc ctccaaaatc ggatctggaa
     2281 gttctgttcc aggggcccct gtccagcggt ctggttccgc gtggttccgg taccgcggcc
     2341 cagccggcca ttttcatgga ccgcctggca agcaaaaaac tgtgtgctga cgacgaatgc
     2401 gtttatacga ttagcctggc aagcgcccaa gaagactaca acgcaccgga ttgccgtttc
     2461 atcaacgtga aaaaaggcca gcaaatctat gtctacagca aactggtgaa agaaaacggc
     2521 gcgggtgaat tttgggccgg ttctgtttat ggcgatggtc aggacgaaat gggcgtggtt
     2581 ggttacttcc cgcgtaatct ggttaaagaa cagcgcgtct atcaagaagc gaccaaagaa
     2641 gtcccgacca cggatattga ctttttctgt gaagcggccg cataataaac cgatacaatt
     2701 aaaggctcct tttggagcct tttttttgga gattttcaac gtgaaaaaat tattattcgc
     2761 aattccttta gttgttcctt tctattctca ctccgctgaa actgttgaaa gttgtttagc
     2821 aaaaccccat acagaaaatt catttactaa cgtctggaaa gacgacaaaa ctttagatcg
     2881 ttacgctaac tatgagggct gtctgtggaa tgctacaggc gttgtagttt gtactggtga
     2941 cgaaactcag tgtctagcta gagtggcggt ggctctggtt ccggtgattt tgattatgaa
     3001 aagatggcaa acgctaataa gggggctatg accgaaaatg ccgatgaaaa cgcgctacag
     3061 tctgacgcta aaggcaaact tgattctgtc gctactgatt acggtgctgc tatcgatggt
     3121 ttcattggtg acgtttccgg ccttgctaat ggtaatggtg ctactggtga ttttgctggc
     3181 tctaattccc aaatggctca agtcggtgac ggtgataatt cacctttaat gaataatttc
     3241 cgtcaatatt taccttccct ccctcaatcg gttgaatgtc gcccttttgt ctttggcgct
     3301 ggtaaaccat atgaattttc tattgattgt gacaaaataa acttattccg tggtgtcttt
     3361 gcgtttcttt tatatgttgc cacctttatg tatgtatttt ctacgtttgc taacatactg
     3421 cgtaataagg agtcttaatc atgccagttc ttttggctag cgccgcccta taccttgtct
     3481 gcctccccgc gttgcgtcgc ggtgcatgga gccgggccac ctcgacctga atggaagccg
     3541 gcggcacctc gctaacggat tcaccactcc aagaattgga gccaatcaat tcttgcggag
     3601 aactgtgaat gcgcaaacca acccttggca gaacatatcc atcgcgtccg ccatctccag
     3661 cagccgcacg cggcgcatct cgggcagcgt tgggtcctgg ccacgggtgc gcatgatcgt
     3721 gctcctgtcg ttgaggaccc ggctaggctg gcggggttgc cttactggtt agcagaatga
     3781 atcaccgata cgcgagcgaa cgtgaagcga ctgctgctgc aaaacgtctg cgacctgagc
     3841 aacaacatga atggtcttcg gtttccgtgt ttcgtaaagt ctggaaacgc ggaagtcagc
     3901 gccctgcacc attatgttcc ggatctgcat cgcaggatgc tgctggctac cctgtggaac
     3961 acctacatct gtattaacga agcgctggca ttgaccctga gtgatttttc tctggtcccg
     4021 ccgcatccat accgccagtt gtttaccctc acaacgttcc agtaaccggg catgttcatc
     4081 atcagtaacc cgtatcgtga gcatcctctc tcgtttcatc ggtatcatta cccccatgaa
     4141 cagaaatccc ccttacacgg aggcatcagt gaccaaacag gaaaaaaccg cccttaacat
     4201 ggcccgcttt atcagaagcc agacattaac gcttctggag aaactcaacg agctggacgc
     4261 ggatgaacag gcagacatct gtgaatcgct tcacgaccac gctgatgagc tttaccgcag
     4321 gatccgcgaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat
     4381 cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata
     4441 gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt
     4501 ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc
     4561 atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa
     4621 agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg
     4681 gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt
     4741 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcgcgga tcctgcctcg
     4801 cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag
     4861 cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg
     4921 gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga tagcggagtg tatactggct
     4981 taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc
     5041 gcacagatgc gtaaggagaa aataccgcat caggcgctct tccgcttcct cgctcactga
     5101 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     5161 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     5221 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     5281 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     5341 aagataccag ggtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     5401 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     5461 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     5521 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     5581 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     5641 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg
     5701 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     5761 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     5821 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     5881 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     5941 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     6001 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     6061 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     6121 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     6181 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     6241 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     6301 gttaatagtt tgcgcaacgt tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg
     6361 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     6421 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     6481 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     6541 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     6601 atgcggcgac cgagttgctc ttgcccggcg tcaacacggg ataataccgc gccacatagc
     6661 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     6721 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     6781 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     6841 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     6901 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     6961 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa
     7021 accattatta tcatgacatt aacctataaa
//
