LOCUS       sequence-192397-        6945 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            CASK-1/1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6945)
  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 6945)
  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_CASK-1/1
FEATURES             Location/Qualifiers
     source          1..6945
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     20..38
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        61..138
                     /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             139..1221
                     /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(158..177)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1464..1469,1470..1485,1486..1492)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1500..1516
                     /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     1505..1527
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1601..1618
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1634..2287
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2245..2267
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2294..2317
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2330..2347
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2603..2650
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4243..4698)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4380..4401)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4592..4611
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4740..4762)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4848..4988
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4903..4922
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5003..5020)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5174..5761)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5932..6723,6724..6792))
                     /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     6555..6574
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6793..6897)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtcttc aagaattccc
       61 gacaccatcg aatggtgcaa aacctttcgc ggtatggcat gatagcgccc ggaagagagt
      121 caattcaggg tggtgaatgt gaaaccagta acgttatacg atgtcgcaga gtatgccggt
      181 gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg
      241 cgggaaaaag tggaagcggc gatggcggag ctgaattaca ttcccaaccg cgtggcacaa
      301 caactggcgg gcaaacagtc gttgctgatt ggcgttgcca cctccagtct ggccctgcac
      361 gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg
      421 gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta aagcggcggt gcacaatctt
      481 ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc tggatgacca ggatgccatt
      541 gctgtggaag ctgcctgcac taatgttccg gcgttatttc ttgatgtctc tgaccagaca
      601 cccatcaaca gtattatttt ctcccatgaa gacggtacgc gactgggcgt ggagcatctg
      661 gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg
      721 cgtctgcgtc tggctggctg gcataaatat ctcactcgca atcaaattca gccgatagcg
      781 gaacgggaag gcgactggag tgccatgtcc ggttttcaac aaaccatgca aatgctgaat
      841 gagggcatcg ttcccactgc gatgctggtt gccaacgatc agatggcgct gggcgcaatg
      901 cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt gggatacgac
      961 gataccgaag acagctcatg ttatatcccg ccgttaacca ccatcaaaca ggattttcgc
     1021 ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag
     1081 ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg
     1141 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     1201 cgactggaaa gcgggcagtg agcgcaacgc aattaatgta agttagctca ctcattaggc
     1261 acaattctca tgtttgacag cttatcatcg actgcacggt gcaccaatgc ttctggcgtc
     1321 aggcagccat cggaagctgt ggtatggctg tgcaggtcgt aaatcactgc ataattcgtg
     1381 tcgctcaagg cgcactcccg ttctggataa tgttttttgc gccgacatca taacggttct
     1441 ggcaaatatt ctgaaatgag ctgttgacaa ttaatcatcg gctcgtataa tgtgtggaat
     1501 tgtgagcgga taacaatttc acacaggaaa cagccagtcc gtttaggtgt tttcacgagc
     1561 acttcaccaa caaggaccat agattatgaa aatcgaagaa caccatcacc atcaccattc
     1621 cagcggtaag cttatgtccc ctatactagg ttattggaaa attaagggcc ttgtgcaacc
     1681 cactcgactt cttttggaat atcttgaaga aaaatatgaa gagcatttgt atgagcgcga
     1741 tgaaggtgat aaatggcgaa acaaaaagtt tgaattgggt ttggagtttc ccaatcttcc
     1801 ttattatatt gatggtgatg ttaaattaac acagtctatg gccatcatac gttatatagc
     1861 tgacaagcac aacatgttgg gtggttgtcc aaaagagcgt gcagagattt caatgcttga
     1921 aggagcggtt ttggatatta gatacggtgt ttcgagaatt gcatatagta aagactttga
     1981 aactctcaaa gttgattttc ttagcaagct acctgaaatg ctgaaaatgt tcgaagatcg
     2041 tttatgtcat aaaacatatt taaatggtga tcatgtaacc catcctgact tcatgttgta
     2101 tgacgctctt gatgttgttt tatacatgga cccaatgtgc ctggatgcgt tcccaaaatt
     2161 agtttgtttt aaaaaacgta ttgaagctat cccacaaatt gataagtact tgaaatccag
     2221 caagtatata gcatggcctt tgcagggctg gcaagccacg tttggtggtg gcgaccatcc
     2281 tccaaaatcg gatctggaag ttctgttcca ggggcccctg tccagcggtc tggttccgcg
     2341 tggttccggt accgcggccc agccggccat ggcccagggc gcgctgattt atgtccgtgc
     2401 ccagtttgaa tatgatccgg ccaaagatga tctgattccg tgcaaagaag ccggtattcg
     2461 ttttcgtgtt ggtgacatta tccagattat ctccaaagat gaccataact ggtggcaggg
     2521 taaactggaa aactccaaaa acggtacggc aggtctgatt ccatcttccg aactgcaaga
     2581 atggcgtgcg gccgcataat aaaccgatac aattaaaggc tccttttgga gccttttttt
     2641 tggagatttt caacgtgaaa aaattattat tcgcaattcc tttagttgtt cctttctatt
     2701 ctcactccgc tgaaactgtt gaaagttgtt tagcaaaacc ccatacagaa aattcattta
     2761 ctaacgtctg gaaagacgac aaaactttag atcgttacgc taactatgag ggctgtctgt
     2821 ggaatgctac aggcgttgta gtttgtactg gtgacgaaac tcagtgtcta gctagagtgg
     2881 cggtggctct ggttccggtg attttgatta tgaaaagatg gcaaacgcta ataagggggc
     2941 tatgaccgaa aatgccgatg aaaacgcgct acagtctgac gctaaaggca aacttgattc
     3001 tgtcgctact gattacggtg ctgctatcga tggtttcatt ggtgacgttt ccggccttgc
     3061 taatggtaat ggtgctactg gtgattttgc tggctctaat tcccaaatgg ctcaagtcgg
     3121 tgacggtgat aattcacctt taatgaataa tttccgtcaa tatttacctt ccctccctca
     3181 atcggttgaa tgtcgccctt ttgtctttgg cgctggtaaa ccatatgaat tttctattga
     3241 ttgtgacaaa ataaacttat tccgtggtgt ctttgcgttt cttttatatg ttgccacctt
     3301 tatgtatgta ttttctacgt ttgctaacat actgcgtaat aaggagtctt aatcatgcca
     3361 gttcttttgg ctagcgccgc cctatacctt gtctgcctcc ccgcgttgcg tcgcggtgca
     3421 tggagccggg ccacctcgac ctgaatggaa gccggcggca cctcgctaac ggattcacca
     3481 ctccaagaat tggagccaat caattcttgc ggagaactgt gaatgcgcaa accaaccctt
     3541 ggcagaacat atccatcgcg tccgccatct ccagcagccg cacgcggcgc atctcgggca
     3601 gcgttgggtc ctggccacgg gtgcgcatga tcgtgctcct gtcgttgagg acccggctag
     3661 gctggcgggg ttgccttact ggttagcaga atgaatcacc gatacgcgag cgaacgtgaa
     3721 gcgactgctg ctgcaaaacg tctgcgacct gagcaacaac atgaatggtc ttcggtttcc
     3781 gtgtttcgta aagtctggaa acgcggaagt cagcgccctg caccattatg ttccggatct
     3841 gcatcgcagg atgctgctgg ctaccctgtg gaacacctac atctgtatta acgaagcgct
     3901 ggcattgacc ctgagtgatt tttctctggt cccgccgcat ccataccgcc agttgtttac
     3961 cctcacaacg ttccagtaac cgggcatgtt catcatcagt aacccgtatc gtgagcatcc
     4021 tctctcgttt catcggtatc attaccccca tgaacagaaa tcccccttac acggaggcat
     4081 cagtgaccaa acaggaaaaa accgccctta acatggcccg ctttatcaga agccagacat
     4141 taacgcttct ggagaaactc aacgagctgg acgcggatga acaggcagac atctgtgaat
     4201 cgcttcacga ccacgctgat gagctttacc gcaggatccg cgaaattgta agcgttaata
     4261 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     4321 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     4381 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     4441 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     4501 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     4561 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     4621 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     4681 cgccgctaca gggcgcgtcg cggatcctgc ctcgcgcgtt tcggtgatga cggtgaaaac
     4741 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     4801 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc
     4861 cagtcacgta gcgatagcgg agtgtatact ggcttaacta tgcggcatca gagcagattg
     4921 tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
     4981 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     5041 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     5101 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     5161 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     5221 caagtcagag gtggcgaaac ccgacaggac tataaagata ccagggtttc cccctggaag
     5281 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5341 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5401 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5461 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5521 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5581 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     5641 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5701 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5761 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5821 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     5881 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     5941 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     6001 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     6061 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     6121 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     6181 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     6241 cattgctgca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     6301 ttcccaacga tcaaggcgag ttacatgatc ccccatattg tgcaaaaaag cggttagctc
     6361 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     6421 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     6481 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     6541 ggcgtcaaca cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     6601 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     6661 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     6721 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     6781 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     6841 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     6901 atttccccga aaagtgccac ctgacgtcta agaaaccatt attat
//
