LOCUS       sequence-488748-        7126 bp    DNA     circular SYN 08-OCT-2025
DEFINITION  Production of His-tagged full-length SHP2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7126)
  AUTHORS   van Vlimmeren AE, Voleti R, Chartier CA, Jiang Z, Karandur D, 
            Humphries PA, Lo WL, Shah NH
  TITLE     The pathogenic T42A mutation in SHP2 rewires the interaction 
            specificity of its N-terminal regulatory domain.
  JOURNAL   Proc Natl Acad Sci U S A. 2024 Jul 23;121(30):e2407159121. doi: 
            10.1073/pnas.2407159121. Epub 2024 Jul 16.
  PUBMED    39012820
REFERENCE   2  (bases 1 to 7126)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Proc Natl 
            Acad Sci U S A."; date: "2024-07-23"; volume: "121(30):e2407159121. 
            doi"; pages: " 10.1073/pnas.2407159121. Epub 2024 Jul 16"
COMMENT     Sequence Label: pET-28b His6-TEV-SHP2-FL(R138Q)
FEATURES             Location/Qualifiers
     source          1..7126
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             9..29
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1861..1878
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1931..1949)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1945..1992
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2029..2484
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2116..2135)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2326..2347
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2576..3391)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2576..3391)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3299..3318
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3383..3402
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3513..4101
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4002..4021
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4255..4272
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4287..4426
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4352..4371)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4512..4534
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4528..4719)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5528..6610)
                     /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     6572..6591
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6611..6688)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6894..6913
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6997..7016
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6997..7015
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    7016..7040
                     /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)."
     RBS             7055..7077
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             7096..7113
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 caacgaccga aaacctttac ttccagggcc atatgacatc gcggagatgg tttcacccaa
       61 atatcactgg tgtggaggca gaaaacctac tgttgacaag aggagttgat ggcagttttt
      121 tggcaaggcc tagtaaaagt aaccctggag acttcacact ttccgttaga agaaatggag
      181 ctgtcaccca catcaagatt cagaacactg gtgattacta tgacctgtat ggaggggaga
      241 aatttgccac tttggctgag ttggtccagt attacatgga acatcacggg caattaaaag
      301 agaagaatgg agatgtcatt gagcttaaat atcctctgaa ctgtgcagat cctacctctg
      361 aaaggtggtt tcatggacat ctctctggga aagaagcaga gaaattatta actgaaaaag
      421 gaaaacatgg tagttttctt gtacaagaga gccagagcca ccctggagat tttgttcttt
      481 ctgtgcgcac tggtgatgac aaaggggaga gcaatgacgg caagtctaaa gtgacccatg
      541 ttatgattcg ctgtcaggaa ctgaaatacg acgttggtgg aggagaacgg tttgattctt
      601 tgacagatct tgtggaacat tataagaaga atcctatggt ggaaacattg ggtacagtac
      661 tacaactcaa gcagcccctt aacacgactc gtataaatgc tgctgaaata gaaagcagag
      721 ttcgagaact aagcaaatta gctgagacca cagataaagt caaacaaggc ttttgggaag
      781 aatttgagac actacaacaa caggagtgca aacttctcta cagccgaaaa gagggtcaaa
      841 ggcaagaaaa caaaaacaaa aatagatata aaaacatcct gccctttgat cataccaggg
      901 ttgtcctaca cgatggtgat cccaatgagc ctgtttcaga ttacatcaat gcaaatatca
      961 tcatgcctga atttgaaacc aagtgcaaca attcaaagcc caaaaagagt tacattgcca
     1021 cacaaggctg cctgcaaaac acggtgaatg acttttggcg gatggtgttc caagaaaact
     1081 cccgagtgat tgtcatgaca acgaaagaag tggagagagg aaagagtaaa tgtgtcaaat
     1141 actggcctga tgagtatgct ctaaaagaat atggcgtcat gcgtgttagg aacgtcaaag
     1201 aaagcgccgc tcatgactat acgctaagag aacttaaact ttcaaaggtt ggacaaggga
     1261 atacggagag aacggtctgg caataccact ttcggacctg gccggaccac ggcgtgccca
     1321 gcgaccctgg gggcgtgctg gacttcctgg aggaggtgca ccataagcag gagagcatca
     1381 tggatgcagg gccggtcgtg gtgcactgca gtgctggaat tggccggaca gggacgttca
     1441 ttgtgattga tattcttatt gacatcatca gagagaaagg tgttgactgc gatattgacg
     1501 ttcccaaaac catccagatg gtgcggtctc agaggtcagg gatggtccag acagaagcac
     1561 agtaccgatt tatctatatg gcggtccagc attatattga aacactacag cgcaggattg
     1621 aagaagagca gaaaagcaag aggaaagggc acgaatatac aaatattaag tattctctag
     1681 cggaccagac gagtggagat cagagccctc tcccgccttg tactccaacg ccaccctgtg
     1741 cagaaatgag agaagacagt gctagagtct atgaaaacgt gggcctgatg caacagcaga
     1801 aaagtttcag atagggatcc gaattcgagc tccgtcgaca agcttgcggc cgcactcgag
     1861 caccaccacc accaccactg agatccggct gctaacaaag cccgaaagga agctgagttg
     1921 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg
     1981 aggggttttt tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta
     2041 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     2101 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     2161 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     2221 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     2281 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     2341 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     2401 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     2461 acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     2521 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgaatt aattcttaga
     2581 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
     2641 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
     2701 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
     2761 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
     2821 ccggtgagaa tggcaaaagt ttatgcattt ctttccagac ttgttcaaca ggccagccat
     2881 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
     2941 gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
     3001 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
     3061 ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcatcatcag
     3121 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
     3181 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
     3241 ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat
     3301 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctag
     3361 agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag
     3421 cagacagttt tattgttcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     3481 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     3541 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     3601 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     3661 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     3721 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     3781 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     3841 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     3901 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     3961 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     4021 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     4081 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     4141 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     4201 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     4261 tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc
     4321 ggtatttcac accgcaatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     4381 gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc
     4441 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     4501 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     4561 gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg
     4621 ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa
     4681 gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg
     4741 atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt
     4801 tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg
     4861 gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga
     4921 tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt
     4981 gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca
     5041 tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat
     5101 cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga
     5161 caggagcacg atcatgcgca cccgtggggc cgccatgccg gcgataatgg cctgcttctc
     5221 gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt gcaagattcc
     5281 gaataccgca agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa
     5341 aatgacccag agcgctgccg gcacctgtcc tacgagttgc atgataaaga agacagtcat
     5401 aagtgcggcg acgatagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc
     5461 tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc
     5521 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     5581 cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca
     5641 ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca
     5701 agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg
     5761 ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa
     5821 cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa
     5881 ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg
     5941 acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat
     6001 atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca
     6061 gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt
     6121 catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg
     6181 gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa
     6241 tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga
     6301 cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt
     6361 taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa
     6421 tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct
     6481 ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca
     6541 ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta
     6601 ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc
     6661 gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc
     6721 tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat
     6781 ggtgcatgca aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc
     6841 acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg
     6901 tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt
     6961 ccggcgtaga ggatcgagat ctcgatcccg cgaaattaat acgactcact ataggggaat
     7021 tgtgagcgga taacaattcc cctctagaaa taattttgtt taactttaag aaggagatat
     7081 accatgggca gcagccatca tcatcatcat cacgattacg atatcc
//
