LOCUS       sequence-65629-e        5450 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 5450)
  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 5450)
  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 Mist-SH2
FEATURES             Location/Qualifiers
     source          1..5450
                     /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"
     CDS             1392..1436
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(1500..1522)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1667..1685)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1753..1857
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1858..1926,1927..2718)
                     /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(2076..2095)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2889..3477
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3378..3397
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3631..3648
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3721..3798
                     /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             3799..4881
                     /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(3818..3837)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    4894..4915
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4930..4935,4936..4953,4954..4960)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4968..4984
                     /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     4992..5008
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4992..5008
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5004..5450,1..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(5024..5041)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5024..5040)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5033..5055)
                     /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 gatgagaagg
      961 atgtctggca gaatgaatgg tacattggag aatacagtcg ccaggcagtg gaagatgtgt
     1021 taatgaaaga gaacaaggat ggtacttttt tggtccgaga ctgctctaca aaatccaagg
     1081 cagaaccata tgttttggtg gtgttttatg ggaacaaggt ctacaatgtg aaaatccgtt
     1141 tcctcgagag caatcaacag tttgccctgg gcacaggact acgaggaaat gagatgtttg
     1201 attctgtgga agacatcatt gaacactaca catattttcc cattctgcta atagatggga
     1261 aagacaaggc tgcacgcagg aaacagtgct acctcaccca gccactgcct ctcgccaggc
     1321 tccttctcac tcagtactcc agccaggcac ttcatgaggg aattcatcgt gacgcaagct
     1381 tgcttggtgg cggtctgaac gacatcttcg aggctcagaa aatcgaatgg cacgaataag
     1441 cggccgcatc gtgactgact gacgatctgc ctcgcgcgtt tcggtgatga cggtgaaaac
     1501 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     1561 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc
     1621 cagtcacgta gcgatagcgg agtgtataat tcttgaagac gaaagggcct cgtgatacgc
     1681 ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt
     1741 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     1801 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg
     1861 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     1921 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     1981 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     2041 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     2101 gttgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     2161 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
     2221 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
     2281 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     2341 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     2401 gcagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
     2461 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
     2521 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
     2581 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     2641 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     2701 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
     2761 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     2821 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     2881 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     2941 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     3001 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     3061 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     3121 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     3181 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     3241 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     3301 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     3361 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     3421 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     3481 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     3541 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     3601 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag
     3661 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg cataaattcc
     3721 gacaccatcg aatggtgcaa aacctttcgc ggtatggcat gatagcgccc ggaagagagt
     3781 caattcaggg tggtgaatgt gaaaccagta acgttatacg atgtcgcaga gtatgccggt
     3841 gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg
     3901 cgggaaaaag tggaagcggc gatggcggag ctgaattaca ttcccaaccg cgtggcacaa
     3961 caactggcgg gcaaacagtc gttgctgatt ggcgttgcca cctccagtct ggccctgcac
     4021 gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg
     4081 gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta aagcggcggt gcacaatctt
     4141 ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc tggatgacca ggatgccatt
     4201 gctgtggaag ctgcctgcac taatgttccg gcgttatttc ttgatgtctc tgaccagaca
     4261 cccatcaaca gtattatttt ctcccatgaa gacggtacgc gactgggcgt ggagcatctg
     4321 gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg
     4381 cgtctgcgtc tggctggctg gcataaatat ctcactcgca atcaaattca gccgatagcg
     4441 gaacgggaag gcgactggag tgccatgtcc ggttttcaac aaaccatgca aatgctgaat
     4501 gagggcatcg ttcccactgc gatgctggtt gccaacgatc agatggcgct gggcgcaatg
     4561 cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt gggatacgac
     4621 gataccgaag acagctcatg ttatatcccg ccgtcaacca ccatcaaaca ggattttcgc
     4681 ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag
     4741 ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg
     4801 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     4861 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc
     4921 accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata
     4981 acaatttcac acaggaaaca gctatgacca tgattacgga ttcactggcc gtcgttttac
     5041 aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc
     5101 ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc
     5161 gcagcctgaa tggcgaatgg cgctttgcct ggtttccggc accagaagcg gtgccggaaa
     5221 gctggctgga gtgcgatctt cctgaggccg atactgtcgt cgtcccctca aactggcaga
     5281 tgcacggtta cgatgcgccc atctacacca acgtaaccta tcccattacg gtcaatccgc
     5341 cgtttgttcc cacggagaat ccgacgggtt gttactcgct cacatttaat gttgatgaaa
     5401 gctggctaca ggaaggccag acgcgaatta tttttgatgg cgttggaatt
//
