LOCUS       sequence-192602-        6900 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            CSK-1/1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6900)
  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 6900)
  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_CSK-1/1
FEATURES             Location/Qualifiers
     source          1..6900
                     /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      2543..2590
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4183..4638)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4320..4341)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4532..4551
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4680..4702)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4788..4928
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4843..4862
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4943..4960)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5114..5701)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5872..6663,6664..6732))
                     /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     6495..6514
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6733..6837)
                     /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 gccaccgaat gcattgccaa atacaacttc catggtacgg cagaacagga
     2401 tctgccgttc tgtaaaggtg atgttctgac cattgtcgcc gttaccaaag atccgaactg
     2461 gtacaaagcc aaaaacaaag ttggtcgtga aggtatcatt ccagccaact atgtccagaa
     2521 acgtgaagcg gccgcataat aaaccgatac aattaaaggc tccttttgga gccttttttt
     2581 tggagatttt caacgtgaaa aaattattat tcgcaattcc tttagttgtt cctttctatt
     2641 ctcactccgc tgaaactgtt gaaagttgtt tagcaaaacc ccatacagaa aattcattta
     2701 ctaacgtctg gaaagacgac aaaactttag atcgttacgc taactatgag ggctgtctgt
     2761 ggaatgctac aggcgttgta gtttgtactg gtgacgaaac tcagtgtcta gctagagtgg
     2821 cggtggctct ggttccggtg attttgatta tgaaaagatg gcaaacgcta ataagggggc
     2881 tatgaccgaa aatgccgatg aaaacgcgct acagtctgac gctaaaggca aacttgattc
     2941 tgtcgctact gattacggtg ctgctatcga tggtttcatt ggtgacgttt ccggccttgc
     3001 taatggtaat ggtgctactg gtgattttgc tggctctaat tcccaaatgg ctcaagtcgg
     3061 tgacggtgat aattcacctt taatgaataa tttccgtcaa tatttacctt ccctccctca
     3121 atcggttgaa tgtcgccctt ttgtctttgg cgctggtaaa ccatatgaat tttctattga
     3181 ttgtgacaaa ataaacttat tccgtggtgt ctttgcgttt cttttatatg ttgccacctt
     3241 tatgtatgta ttttctacgt ttgctaacat actgcgtaat aaggagtctt aatcatgcca
     3301 gttcttttgg ctagcgccgc cctatacctt gtctgcctcc ccgcgttgcg tcgcggtgca
     3361 tggagccggg ccacctcgac ctgaatggaa gccggcggca cctcgctaac ggattcacca
     3421 ctccaagaat tggagccaat caattcttgc ggagaactgt gaatgcgcaa accaaccctt
     3481 ggcagaacat atccatcgcg tccgccatct ccagcagccg cacgcggcgc atctcgggca
     3541 gcgttgggtc ctggccacgg gtgcgcatga tcgtgctcct gtcgttgagg acccggctag
     3601 gctggcgggg ttgccttact ggttagcaga atgaatcacc gatacgcgag cgaacgtgaa
     3661 gcgactgctg ctgcaaaacg tctgcgacct gagcaacaac atgaatggtc ttcggtttcc
     3721 gtgtttcgta aagtctggaa acgcggaagt cagcgccctg caccattatg ttccggatct
     3781 gcatcgcagg atgctgctgg ctaccctgtg gaacacctac atctgtatta acgaagcgct
     3841 ggcattgacc ctgagtgatt tttctctggt cccgccgcat ccataccgcc agttgtttac
     3901 cctcacaacg ttccagtaac cgggcatgtt catcatcagt aacccgtatc gtgagcatcc
     3961 tctctcgttt catcggtatc attaccccca tgaacagaaa tcccccttac acggaggcat
     4021 cagtgaccaa acaggaaaaa accgccctta acatggcccg ctttatcaga agccagacat
     4081 taacgcttct ggagaaactc aacgagctgg acgcggatga acaggcagac atctgtgaat
     4141 cgcttcacga ccacgctgat gagctttacc gcaggatccg cgaaattgta agcgttaata
     4201 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     4261 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     4321 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     4381 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     4441 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     4501 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     4561 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     4621 cgccgctaca gggcgcgtcg cggatcctgc ctcgcgcgtt tcggtgatga cggtgaaaac
     4681 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     4741 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc
     4801 cagtcacgta gcgatagcgg agtgtatact ggcttaacta tgcggcatca gagcagattg
     4861 tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
     4921 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     4981 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     5041 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     5101 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     5161 caagtcagag gtggcgaaac ccgacaggac tataaagata ccagggtttc cccctggaag
     5221 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5281 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5341 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5401 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5461 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5521 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     5581 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5641 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5701 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5761 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     5821 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     5881 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     5941 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     6001 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     6061 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     6121 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     6181 cattgctgca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     6241 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     6301 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     6361 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     6421 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     6481 ggcgtcaaca cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     6541 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     6601 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     6661 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     6721 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     6781 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     6841 atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta
//
