LOCUS       sequence-191792-        6963 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            SH3RF2-1/3-SV1/2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6963)
  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 6963)
  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_SH3RF2-1/3-SV1/2
FEATURES             Location/Qualifiers
     source          1..6963
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     27..45
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        68..145
                     /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             146..1228
                     /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(165..184)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1471..1476,1477..1492,1493..1499)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1507..1523
                     /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     1512..1534
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1608..1625
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1641..2294
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2252..2274
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2301..2324
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2337..2354
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2628..2675
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4268..4723)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4405..4426)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4617..4636
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4765..4787)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4873..5013
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4928..4947
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5028..5045)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5199..5786)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5957..6748,6749..6817))
                     /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     6580..6599
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6818..6922)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcaag
       61 aattcccgac accatcgaat ggtgcaaaac ctttcgcggt atggcatgat agcgcccgga
      121 agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg tcgcagagta
      181 tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc acgtttctgc
      241 gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc ccaaccgcgt
      301 ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct ccagtctggc
      361 cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc
      421 cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca
      481 caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg atgaccagga
      541 tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg atgtctctga
      601 ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac tgggcgtgga
      661 gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt
      721 ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc
      781 gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat
      841 gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg
      901 cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct cggtagtggg
      961 atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca tcaaacagga
     1021 ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc
     1081 ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc
     1141 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     1201 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt tagctcactc
     1261 attaggcaca attctcatgt ttgacagctt atcatcgact gcacggtgca ccaatgcttc
     1321 tggcgtcagg cagccatcgg aagctgtggt atggctgtgc aggtcgtaaa tcactgcata
     1381 attcgtgtcg ctcaaggcgc actcccgttc tggataatgt tttttgcgcc gacatcataa
     1441 cggttctggc aaatattctg aaatgagctg ttgacaatta atcatcggct cgtataatgt
     1501 gtggaattgt gagcggataa caatttcaca caggaaacag ccagtccgtt taggtgtttt
     1561 cacgagcact tcaccaacaa ggaccataga ttatgaaaat cgaagaacac catcaccatc
     1621 accattccag cggtaagctt atgtccccta tactaggtta ttggaaaatt aagggccttg
     1681 tgcaacccac tcgacttctt ttggaatatc ttgaagaaaa atatgaagag catttgtatg
     1741 agcgcgatga aggtgataaa tggcgaaaca aaaagtttga attgggtttg gagtttccca
     1801 atcttcctta ttatattgat ggtgatgtta aattaacaca gtctatggcc atcatacgtt
     1861 atatagctga caagcacaac atgttgggtg gttgtccaaa agagcgtgca gagatttcaa
     1921 tgcttgaagg agcggttttg gatattagat acggtgtttc gagaattgca tatagtaaag
     1981 actttgaaac tctcaaagtt gattttctta gcaagctacc tgaaatgctg aaaatgttcg
     2041 aagatcgttt atgtcataaa acatatttaa atggtgatca tgtaacccat cctgacttca
     2101 tgttgtatga cgctcttgat gttgttttat acatggaccc aatgtgcctg gatgcgttcc
     2161 caaaattagt ttgttttaaa aaacgtattg aagctatccc acaaattgat aagtacttga
     2221 aatccagcaa gtatatagca tggcctttgc agggctggca agccacgttt ggtggtggcg
     2281 accatcctcc aaaatcggat ctggaagttc tgttccaggg gcccctgtcc agcggtctgg
     2341 ttccgcgtgg ttccggtacc gcggcccagc cggccagccc gttccgtctg gttccgaatg
     2401 tccgtattca tatggatggt gtcccgcgtg ccaaagccct gtgtaactac cgtggtcaaa
     2461 acccgggcga tctgcgtttt aacaaaggtg acattatcct gctgcgtcgc cagctggatg
     2521 aaaattggta tcaaggcgaa attaacggca tcagtggtaa tttcccggcg agctctgtgg
     2581 aagttattaa acagctgccg caaccgccgc cggcggccgc ataataaacc gatacaatta
     2641 aaggctcctt ttggagcctt ttttttggag attttcaacg tgaaaaaatt attattcgca
     2701 attcctttag ttgttccttt ctattctcac tccgctgaaa ctgttgaaag ttgtttagca
     2761 aaaccccata cagaaaattc atttactaac gtctggaaag acgacaaaac tttagatcgt
     2821 tacgctaact atgagggctg tctgtggaat gctacaggcg ttgtagtttg tactggtgac
     2881 gaaactcagt gtctagctag agtggcggtg gctctggttc cggtgatttt gattatgaaa
     2941 agatggcaaa cgctaataag ggggctatga ccgaaaatgc cgatgaaaac gcgctacagt
     3001 ctgacgctaa aggcaaactt gattctgtcg ctactgatta cggtgctgct atcgatggtt
     3061 tcattggtga cgtttccggc cttgctaatg gtaatggtgc tactggtgat tttgctggct
     3121 ctaattccca aatggctcaa gtcggtgacg gtgataattc acctttaatg aataatttcc
     3181 gtcaatattt accttccctc cctcaatcgg ttgaatgtcg cccttttgtc tttggcgctg
     3241 gtaaaccata tgaattttct attgattgtg acaaaataaa cttattccgt ggtgtctttg
     3301 cgtttctttt atatgttgcc acctttatgt atgtattttc tacgtttgct aacatactgc
     3361 gtaataagga gtcttaatca tgccagttct tttggctagc gccgccctat accttgtctg
     3421 cctccccgcg ttgcgtcgcg gtgcatggag ccgggccacc tcgacctgaa tggaagccgg
     3481 cggcacctcg ctaacggatt caccactcca agaattggag ccaatcaatt cttgcggaga
     3541 actgtgaatg cgcaaaccaa cccttggcag aacatatcca tcgcgtccgc catctccagc
     3601 agccgcacgc ggcgcatctc gggcagcgtt gggtcctggc cacgggtgcg catgatcgtg
     3661 ctcctgtcgt tgaggacccg gctaggctgg cggggttgcc ttactggtta gcagaatgaa
     3721 tcaccgatac gcgagcgaac gtgaagcgac tgctgctgca aaacgtctgc gacctgagca
     3781 acaacatgaa tggtcttcgg tttccgtgtt tcgtaaagtc tggaaacgcg gaagtcagcg
     3841 ccctgcacca ttatgttccg gatctgcatc gcaggatgct gctggctacc ctgtggaaca
     3901 cctacatctg tattaacgaa gcgctggcat tgaccctgag tgatttttct ctggtcccgc
     3961 cgcatccata ccgccagttg tttaccctca caacgttcca gtaaccgggc atgttcatca
     4021 tcagtaaccc gtatcgtgag catcctctct cgtttcatcg gtatcattac ccccatgaac
     4081 agaaatcccc cttacacgga ggcatcagtg accaaacagg aaaaaaccgc ccttaacatg
     4141 gcccgcttta tcagaagcca gacattaacg cttctggaga aactcaacga gctggacgcg
     4201 gatgaacagg cagacatctg tgaatcgctt cacgaccacg ctgatgagct ttaccgcagg
     4261 atccgcgaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     4321 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     4381 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     4441 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     4501 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     4561 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     4621 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     4681 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcgcggat cctgcctcgc
     4741 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     4801 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     4861 cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat agcggagtgt atactggctt
     4921 aactatgcgg catcagagca gattgtactg agagtgcacc atatgcggtg tgaaataccg
     4981 cacagatgcg taaggagaaa ataccgcatc aggcgctctt ccgcttcctc gctcactgac
     5041 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     5101 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     5161 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     5221 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     5281 agataccagg gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     5341 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     5401 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     5461 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     5521 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     5581 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga
     5641 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     5701 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     5761 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     5821 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5881 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5941 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     6001 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     6061 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     6121 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     6181 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     6241 ttaatagttt gcgcaacgtt gttgccattg ctgcaggcat cgtggtgtca cgctcgtcgt
     6301 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6361 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     6421 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6481 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6541 tgcggcgacc gagttgctct tgcccggcgt caacacggga taataccgcg ccacatagca
     6601 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6661 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6721 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6781 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6841 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6901 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     6961 cca
//
