LOCUS       sequence-65559-e        5264 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5264)
  AUTHORS   Machida K, Thompson CM, Dierck K, Jablonowski K, Karkkainen S, Liu 
            B, Zhang H, Nash PD, Newman DK, Nollau P, Pawson T, Renkema GH, 
            Saksela K, Schiller MR, Shin DG, Mayer BJ
  TITLE     High-throughput phosphotyrosine profiling using SH2 domains.
  JOURNAL   Mol Cell. 2007 Jun 22;26(6):899-915.
  PUBMED    17588523
REFERENCE   2  (bases 1 to 5264)
  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"; journalName: "Mol Cell.";
            date: "2007-06-22"; volume: "26(6)"; pages: "899-915"
COMMENT     Sequence Label: pGEX CrkL-SH2
FEATURES             Location/Qualifiers
     source          1..5264
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..941
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1314..1336)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1481..1499)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1567..1671
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1672..1740,1741..2532)
                     /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     complement(1890..1909)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2703..3291
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3192..3211
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3445..3462
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3535..3612
                     /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             3613..4695
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3632..3651)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    4708..4729
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4744..4749,4750..4767,4768..4774)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4782..4798
                     /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     4806..4822
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4806..4822
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4818..5264,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4838..4855)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4838..4854)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4847..4869)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gaagttctgt tccaggggcc cctgggatcc tcggaccgct
      961 ccgcctggta tatggggccg gtgtctcgcc aggaggcgca gacccggctc cagggccagc
     1021 gccacggtat gttcctcgtc cgcgattctt ccacctgccc tggggactat gtgctgtcgg
     1081 tgtccgagaa ctcgcgggtc tcccactaca tcatcaactc gctgcccaac cgccgtttta
     1141 agatcgggga ccaggaattt gaccatttgc cggccctgct ggagttttac aagatccact
     1201 acctggacac caccaccctc atcgagcctg cgcccaggta tccaagccca taggcggccg
     1261 catcgtgact gactgacgat ctgcctcgcg cgtttcggtg atgacggtga aaacctctga
     1321 cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa
     1381 gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat gacccagtca
     1441 cgtagcgata gcggagtgta taattcttga agacgaaagg gcctcgtgat acgcctattt
     1501 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga
     1561 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     1621 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     1681 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     1741 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     1801 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     1861 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtgttgac
     1921 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     1981 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     2041 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     2101 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     2161 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgcagca
     2221 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     2281 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     2341 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     2401 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     2461 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     2521 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     2581 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     2641 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     2701 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     2761 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     2821 ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac
     2881 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     2941 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     3001 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     3061 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     3121 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     3181 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     3241 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     3301 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     3361 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
     3421 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg
     3481 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcataaa ttccgacacc
     3541 atcgaatggt gcaaaacctt tcgcggtatg gcatgatagc gcccggaaga gagtcaattc
     3601 agggtggtga atgtgaaacc agtaacgtta tacgatgtcg cagagtatgc cggtgtctct
     3661 tatcagaccg tttcccgcgt ggtgaaccag gccagccacg tttctgcgaa aacgcgggaa
     3721 aaagtggaag cggcgatggc ggagctgaat tacattccca accgcgtggc acaacaactg
     3781 gcgggcaaac agtcgttgct gattggcgtt gccacctcca gtctggccct gcacgcgccg
     3841 tcgcaaattg tcgcggcgat taaatctcgc gccgatcaac tgggtgccag cgtggtggtg
     3901 tcgatggtag aacgaagcgg cgtcgaagcc tgtaaagcgg cggtgcacaa tcttctcgcg
     3961 caacgcgtca gtgggctgat cattaactat ccgctggatg accaggatgc cattgctgtg
     4021 gaagctgcct gcactaatgt tccggcgtta tttcttgatg tctctgacca gacacccatc
     4081 aacagtatta ttttctccca tgaagacggt acgcgactgg gcgtggagca tctggtcgca
     4141 ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa gttctgtctc ggcgcgtctg
     4201 cgtctggctg gctggcataa atatctcact cgcaatcaaa ttcagccgat agcggaacgg
     4261 gaaggcgact ggagtgccat gtccggtttt caacaaacca tgcaaatgct gaatgagggc
     4321 atcgttccca ctgcgatgct ggttgccaac gatcagatgg cgctgggcgc aatgcgcgcc
     4381 attaccgagt ccgggctgcg cgttggtgcg gatatctcgg tagtgggata cgacgatacc
     4441 gaagacagct catgttatat cccgccgtca accaccatca aacaggattt tcgcctgctg
     4501 gggcaaacca gcgtggaccg cttgctgcaa ctctctcagg gccaggcggt gaagggcaat
     4561 cagctgttgc ccgtctcact ggtgaaaaga aaaaccaccc tggcgcccaa tacgcaaacc
     4621 gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg
     4681 gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca
     4741 ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
     4801 tcacacagga aacagctatg accatgatta cggattcact ggccgtcgtt ttacaacgtc
     4861 gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat ccccctttcg
     4921 ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc
     4981 tgaatggcga atggcgcttt gcctggtttc cggcaccaga agcggtgccg gaaagctggc
     5041 tggagtgcga tcttcctgag gccgatactg tcgtcgtccc ctcaaactgg cagatgcacg
     5101 gttacgatgc gcccatctac accaacgtaa cctatcccat tacggtcaat ccgccgtttg
     5161 ttcccacgga gaatccgacg ggttgttact cgctcacatt taatgttgat gaaagctggc
     5221 tacaggaagg ccagacgcga attatttttg atggcgttgg aatt
//
