LOCUS       sequence-65599-e        5344 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 5344)
  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 5344)
  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 Grb14-SH2
FEATURES             Location/Qualifiers
     source          1..5344
                     /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(1394..1416)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1561..1579)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1647..1751
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1752..1820,1821..2612)
                     /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(1970..1989)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2783..3371
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3272..3291
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3525..3542
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3615..3692
                     /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             3693..4775
                     /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(3712..3731)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    4788..4809
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4824..4829,4830..4847,4848..4854)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4862..4878
                     /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     4886..4902
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4886..4902
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4898..5344,1..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(4918..4935)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4918..4934)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4927..4949)
                     /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 gggcgcgcca
      961 tgtctgccac aaacatggct atccaccggt cccagccatg gtttcaccac aaaatttcta
     1021 gagatgaggc tcagcgattg attattcagc aaggacttgt ggatggagtt ttcttggtac
     1081 gggatagtca gagtaacccc aaaactttcg tactgtcaat gagtcatgga caaaaaataa
     1141 agcactttca aattatacca gtagaagatg acggtgaaat gttccacaca ctggatgatg
     1201 gccacacaag atttacagat ctaatacagc tggtggagtt ctatcaactc aataagggcg
     1261 ttcttccttg caagttgaaa cattattgtg ctaggattgc tctcttaatt aacgaattcc
     1321 cgggtcgact cgagcggccg catcgtgact gactgacgat ctgcctcgcg cgtttcggtg
     1381 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
     1441 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     1501 gcgcagccat gacccagtca cgtagcgata gcggagtgta taattcttga agacgaaagg
     1561 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     1621 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     1681 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     1741 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     1801 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     1861 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     1921 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     1981 cggtattatc ccgtgttgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     2041 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     2101 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     2161 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     2221 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     2281 acaccacgat gcctgcagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     2341 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     2401 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     2461 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     2521 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     2581 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     2641 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     2701 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     2761 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     2821 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     2881 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     2941 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     3001 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     3061 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     3121 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     3181 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     3241 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     3301 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     3361 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     3421 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     3481 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     3541 aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     3601 accgcataaa ttccgacacc atcgaatggt gcaaaacctt tcgcggtatg gcatgatagc
     3661 gcccggaaga gagtcaattc agggtggtga atgtgaaacc agtaacgtta tacgatgtcg
     3721 cagagtatgc cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag gccagccacg
     3781 tttctgcgaa aacgcgggaa aaagtggaag cggcgatggc ggagctgaat tacattccca
     3841 accgcgtggc acaacaactg gcgggcaaac agtcgttgct gattggcgtt gccacctcca
     3901 gtctggccct gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc gccgatcaac
     3961 tgggtgccag cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc tgtaaagcgg
     4021 cggtgcacaa tcttctcgcg caacgcgtca gtgggctgat cattaactat ccgctggatg
     4081 accaggatgc cattgctgtg gaagctgcct gcactaatgt tccggcgtta tttcttgatg
     4141 tctctgacca gacacccatc aacagtatta ttttctccca tgaagacggt acgcgactgg
     4201 gcgtggagca tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa
     4261 gttctgtctc ggcgcgtctg cgtctggctg gctggcataa atatctcact cgcaatcaaa
     4321 ttcagccgat agcggaacgg gaaggcgact ggagtgccat gtccggtttt caacaaacca
     4381 tgcaaatgct gaatgagggc atcgttccca ctgcgatgct ggttgccaac gatcagatgg
     4441 cgctgggcgc aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg gatatctcgg
     4501 tagtgggata cgacgatacc gaagacagct catgttatat cccgccgtca accaccatca
     4561 aacaggattt tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa ctctctcagg
     4621 gccaggcggt gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga aaaaccaccc
     4681 tggcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
     4741 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag
     4801 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga
     4861 attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta cggattcact
     4921 ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct
     4981 tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc
     5041 ttcccaacag ttgcgcagcc tgaatggcga atggcgcttt gcctggtttc cggcaccaga
     5101 agcggtgccg gaaagctggc tggagtgcga tcttcctgag gccgatactg tcgtcgtccc
     5161 ctcaaactgg cagatgcacg gttacgatgc gcccatctac accaacgtaa cctatcccat
     5221 tacggtcaat ccgccgtttg ttcccacgga gaatccgacg ggttgttact cgctcacatt
     5281 taatgttgat gaaagctggc tacaggaagg ccagacgcga attatttttg atggcgttgg
     5341 aatt
//
