LOCUS       sequence-337604-        8867 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Two-hybrid system that confers spectinomycin resistance in response 
            to substrate phosphorylation. Constitutive expression of c-src 
            kinase, CDC37, and SHP1 (R459M).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8867)
  TITLE     Fox lab plasmids
REFERENCE   2  (bases 1 to 8867)
  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"
COMMENT     Sequence Label: pB2H6*
FEATURES             Location/Qualifiers
     source          1..8867
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(21..39)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        107..211
                     /gene="bla"
                     /label=AmpR promoter
     CDS             212..1027
                     /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             212..1027
                     /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     complement(285..304)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     complement(1073..1090)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1243..1329
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1421..1448
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     CDS             2280..2303
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2623..2895
                     /codon_start=1
                     /gene="E. coli rpoZ"
                     /product="DNA-directed RNA polymerase subunit omega"
                     /label=RNAP omega
                     /note="promotes RNA polymerase assembly"
                     /translation="MARVTVQDAVEKIGNRFDLVLVAARRARQMQVGGKDPLVPEENDK
                     TTVIALREIEEGLINNQILDVRERQEQQEQEAAELQAVTAIAEGRR"
     RBS             5163..5174
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             complement(6167..6958)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      7647..7869
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             7917..8867
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVKIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
ORIGIN
        1 acggcctatt tggcctaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
       61 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
      121 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
      181 aatgcttcaa taatattgaa aaaggaagag tatgagccat attcaacggg aaacgtcttg
      241 ctctaggccg cgattaaatt ccaacatgga tgctgattta tatgggtata aatgggctcg
      301 cgataatgtc gggcaatcag gtgcgacaat ctatcgattg tatgggaagc ccgatgcgcc
      361 agagttgttt ctgaaacatg gcaaaggtag cgttgccaat gatgttacag atgagatggt
      421 cagactaaac tggctgacgg aatttatgcc tcttccgacc atcaagcatt ttatccgtac
      481 tcctgatgat gcatggttac tcaccactgc gatccccggg aaaacagcat tccaggtatt
      541 agaagaatat cctgattcag gtgaaaatat tgttgatgcg ctggcagtgt tcctgcgccg
      601 gttgcattcg attcctgttt gtaattgtcc ttttaacagc gatcgcgtat ttcgtctcgc
      661 tcaggcgcaa tcacgaatga ataacggttt ggttgatgcg agtgattttg atgacgagcg
      721 taatggctgg cctgttgaac aagtctggaa agaaatgcat aaacttttgc cattctcacc
      781 ggattcagtc gtcactcatg gtgatttctc acttgataac cttatttttg acgaggggaa
      841 attaataggt tgtattgatg ttggacgagt cggaatcgca gaccgatacc aggatcttgc
      901 catcctatgg aactgcctcg gtgagttttc tccttcatta cagaaacggc tttttcaaaa
      961 atatggtatt gataatcctg atatgaataa attgcagttt catttgatgc tcgatgagtt
     1021 tttctaaacg ccatgggcat ggctgttttg gcggatgaga gaagattttc agcctgatac
     1081 agattaaatc agaacgcaga agcggtctga taaaacagaa tttgcctggc gacagtagcg
     1141 cggtggtccc acctgacccc atgccgaact cagaagtgaa acgccgtagc gccgatggta
     1201 gtgtggggtc tccccatgcg agagtaggga actgccaggc atcaaataaa acgaaaggct
     1261 cagtcgaaag actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt
     1321 aggacaaatc cgccgggagc ggatttgaac gttgcgaagc aacggcccgg agggtggcgg
     1381 gcaggacgcc cgccataaac tgccaggcat caaattaagc agaaggccat cctgacggat
     1441 ggcctttttg cgtttctaca aactcgctga ggattctaga gcacagctaa caccacgtcg
     1501 tccctatctg ctgccctagg tctatgagtg gttgctggat aactttacgg gcatgcataa
     1561 ggctcggtat ctatattcag ggagaccaca acggtttccc tctacaaata attttgttta
     1621 acttttacta gagtaaggag gaaaaaaaaa tgagtatcag cagcagggta aaaagcaaaa
     1681 gaattcagct tggacttaac caggctgaac ttgctcaaaa ggtggggact acccagcagt
     1741 ctatagagca gctcgaaaac ggtaaaacta agcgaccacg ctttttacca gaacttgcgt
     1801 cagctcttgg cgtaagtgtt gactggctgc tcaatggcac ctctgattcg aatgttagat
     1861 ttgttgggca cgttgagccc aaagggaaat atccattgat tagcatggtt agagctcgtt
     1921 cgtggtgtga agcttgtgaa ccctacgata tcaaggacat tgatgaatgg tatgacagtg
     1981 acgttaactt attaggcaat ggattctggc tgaaggttga aggtgattcc atgacctcac
     2041 ctgtaggtca aagcatccct gaaggtcata tggtgttagt agatactgga cgggagccag
     2101 tgaatggaag ccttgttgta gccaaactga ctgacgcgaa cgaagcaaca ttcaagaaac
     2161 tggtcataga tggcggtcag aagtacctga aaggcctgaa tccttcatgg cctatgactc
     2221 ctatcaacgg aaactgcaag attatcggtg ttgtcgtgga agcgagggta aaattcgtag
     2281 actacaagga cgacgatgac aagtggtatt ttgggaagat cactcgtcgg gagtccgagc
     2341 ggctgctgct caaccccgaa aacccccggg gaaccttctt ggtccgggag agcgagacgg
     2401 taaaaggtgc ctatgccctc tccgtttctg actttgacaa cgccaagggg ctcaatgtga
     2461 aacactacct gatccgcaag ctggacagcg gcggcttcta catcacctca cgcacacagt
     2521 tcagcagcct gcagcagctg gtggcctact actccaaaca tgctgatggc ttgtgccacc
     2581 gcctgaccaa cgtctgctaa gtgcagtaag gaggaaaaaa aaatggcacg cgtaactgtt
     2641 caggacgctg tagagaaaat tggtaaccgt tttgacctgg tactggtcgc cgcgcgtcgc
     2701 gctcgtcaga tgcaggtagg cggaaaggat ccgctggtac cggaagaaaa cgataaaacc
     2761 actgtaatcg cgctgcgcga aatcgaagaa ggtctgatca acaaccagat cctcgacgtt
     2821 cgcgaacgcc aggaacagca agagcaggaa gccgctgaat tacaagccgt taccgctatt
     2881 gctgaaggtc gtcgtgcagc tgcggaaccg cagtatgaag aaattccgat ttatctgtaa
     2941 taattctaga gcacagctaa caccacgtcg tccctatctg ctgccctagg tctatgagtg
     3001 gttgctggat aactttacgg gcatgcataa ggctcgtata atatattcag ggagaccaca
     3061 acggtttccc tctacaaata attttgttta acttttacta gagcgaaaaa aagtaaggcg
     3121 gtaatccatg ggctccaagc cgcagactca gggcctggcc aaggatgcct gggagatccc
     3181 tcgggagtcg ctgcggctgg aggtcaagct gggccagggc tgctttggcg aggtgtggat
     3241 ggggacctgg aacggtacca ccagggtggc catcaaaacc ctgaagcctg gcacgatgtc
     3301 tccagaggcc ttcctgcagg aggcccaggt catgaagaag ctgaggcatg agaagctggt
     3361 gcagttgtat gctgtggttt cagaggagcc catttacatc gtcacggagt acatgagcaa
     3421 ggggagtttg ctggactttc tcaaggggga gacaggcaag tacctgcggc tgcctcagct
     3481 ggtggacatg gctgctcaga tcgcctcagg catggcgtac gtggagcgga tgaactacgt
     3541 ccaccgggac cttcgtgcag ccaacatcct ggtgggagag aacctggtgt gcaaagtggc
     3601 cgactttggg ctggctcggc tcattgaaga caatgagtac acggcgcggc aaggtgccaa
     3661 attccccatc aagtggacgg ctccagaagc tgccctctat ggccgcttca ccatcaagtc
     3721 ggacgtgtgg tccttcggga tcctgctgac tgagctcacc acaaagggac gggtgcccta
     3781 ccctgggatg gtgaaccgcg aggtgctgga ccaggtggag cggggctacc ggatgccctg
     3841 cccgccggag tgtcccgagt ccctgcacga cctcatgtgc cagtgctggc ggaaggagcc
     3901 tgaggagcgg cccaccttcg agtacctgca ggccttcctg gaggactact tcacgtccac
     3961 cgagccccag taccagcccg gggagaacct ctaagctagc tttaagaagg agatatacca
     4021 tggtggacta cagcgtgtgg gaccacattg aggtgtctga tgatgaagac gagacgcacc
     4081 ccaacatcga cacggccagt ctcttccgct ggcggcatca ggcccgggtg gaacgcatgg
     4141 agcagttcca gaaggagaag gaggaactgg acaggggctg ccgcgagtgc aagcgcaagg
     4201 tggccgagtg ccagaggaaa ctgaaggagc tggaggtggc cgagggcggc aaggcagagc
     4261 tggagcgcct gcaggccgag gcacagcagc tgcgcaagga ggagcggagc tgggagcaga
     4321 agctggagga gatgcgcaag aaggagaaga gcatgccctg gaacgtggac acgctcagca
     4381 aagacggctt cagcaagagc atggtaaata ccaagcccga gaagacggag gaggactcag
     4441 aggaggtgag ggagcagaaa cacaagacct tcgtggaaaa atacgagaaa cagatcaagc
     4501 actttggcat gcttcgccgc tgggatgaca gccaaaagta cctgtcagac aacgtccacc
     4561 tggtgtgcga ggagacagcc aattacctgg tcatttggtg cattgaccta gaggtggagg
     4621 agaaatgtgc actcatggag caggtggccc accagacaat cgtcatgcaa tttatcctgg
     4681 agctggccaa gagcctaaag gtggaccccc gggcctgctt ccggcagttc ttcactaaga
     4741 ttaagacagc cgatcgccag tacatggagg gcttcaacga cgagctggaa gccttcaagg
     4801 agcgtgtgcg gggccgtgcc aagctgcgca tcgagaaggc catgaaggag tacgaggagg
     4861 aggagcgcaa gaagcggctc ggccccggcg gcctggaccc cgtcgaggtc tacgagtccc
     4921 tccctgagga actccagaag tgcttcgatg tgaaggacgt gcagatgctg caggacgcca
     4981 tcagcaagat ggaccccacc gacgcaaagt accacatgca gcgctgcatt gactctggcc
     5041 tctgggtccc caactctaag gccagcgagg ccaaggaggg agaggaggca ggtcctgggg
     5101 acccattact ggaagctgtt cccaagacgg gcgatgagaa ggatgtcagt gtgtaagtgc
     5161 agaaagagga gaaatactag atggctgaca ttgagaaccg agtgttggaa ctgaacaaga
     5221 agcaggagtc cgaggataca gccaaggctg gcttctggga ggagtttgag agtttgcaga
     5281 agcaggaggt gaagaacttg caccagcgtc tggaagggca gcggccagag aacaagggca
     5341 agaaccgcta caagaacatt ctcccctttg accacagccg agtgatcctg cagggacggg
     5401 acagtaacat ccccgggtcc gactacatca atgccaacta catcaagaac cagctgctag
     5461 gccctgatga gaacgctaag acctacatcg ccagccaggg ctgtctggag gccacggtca
     5521 atgacttctg gcagatggcg tggcaggaga acagccgtgt catcgtcatg accacccgag
     5581 aggtggagaa aggccggaac aaatgcgtcc catactggcc cgaggtgggc atgcagcgtg
     5641 cttatgggcc ctactctgtg accaactgcg gggagcatga cacaaccgaa tacaaactcc
     5701 gtaccttaca ggtctccccg ctggacaatg gagacctgat tcgggagatc tggcattacc
     5761 agtacctgag ctggcccgac catggggtcc ccagtgagcc tgggggtgtc ctcagcttcc
     5821 tggaccagat caaccagcgg caggaaagtc tgcctcacgc agggcccatc atcgtgcact
     5881 gcagcgccgg catcggcatg acaggcacca tcattgtcat cgacatgctc atggagaaca
     5941 tctccaccaa gggcctggac tgtgacattg acatccagaa gaccatccag atggtgcggg
     6001 cgcagcgctc gggcatggtg cagacggagg cgcagtacaa gttcatctac gtggccatcg
     6061 cccagttcat tgaaaccact aagaagaagc tgtgacagtc agtagggccc taaaaaaaaa
     6121 ccccgcccct gacaggggtg gggttttttg cgtacattgg ctccgttcat ttgccgacta
     6181 ccttggtgat ctcgcctttc acgtagtgga caaattcttc caactgatct gcgcgcgagg
     6241 ccaagcgatc ttcttcttgt ccaagataag cctgtctagc ttcaagtatg acgggctgat
     6301 actgggccgg caggcgctcc attgcccagt cggcagcgac atccttcggc gcgattttgc
     6361 cggttactgc gctgtaccaa atgcgggaca acgtaagcac tacatttcgc tcatcgccag
     6421 cccagtcggg cggcgagttc catagcgtta aggtttcatt tagcgcctca aatagatcct
     6481 gttcaggaac cggatcaaag agttcctccg ccgctggacc taccaaggca acgctatgtt
     6541 ctcttgcttt tgtcagcaag atagccagat caatgtcgat cgtggctggc tcgaagatac
     6601 ctgcaagaat gtcattgcgc tgccattctc caaattgcag ttcgcgctta gctggataac
     6661 gccacggaat gatgtcgtcg tgcacaacaa tggtgacttc tacagcgcgg agaatctcgc
     6721 tctctccagg ggaagccgaa gtttccaaaa ggtcgttgat caaagctcgc cgcgttgttt
     6781 catcaagcct tacggtcacc gtaaccagca aatcaatatc actgtgtggc ttcaggccgc
     6841 catccactgc ggagccgtac aaatgtacgg ccagcaacgt cggttcgaga tggcgctcga
     6901 tgacgccaac tacctctgat agttgagtcg atacttcggc gatcaccgct tccctcattt
     6961 ttttttcctc cttactgcac ctgcaggtcg gtcgacacaa catacgcgcc ggaagcttta
     7021 cgagtaaagc ccggggtgcc taatgaacaa actttcttgt attttcatac aatgtatctt
     7081 gttttagggt ccctaagtct ccgcagcgca acgaaaggcc cagtctttcg actgagcctt
     7141 tcgttgcatt tgcccggatg tcctcttgac cgtacgacgg gtaagcctgt tgatgatacc
     7201 gctgccttac tgggtgcatt agccagtctg aatgacctgt cacgggataa tccgaagtgg
     7261 tcagactgga aaatcagagg gcaggaactg ctgaacagca aaaagtcaga tagcaccaca
     7321 tagcagaccc gccataaaac gccctgagaa gcccgtgacg ggcttttctt gtattatggg
     7381 tagtttcctt gcatgaatcc ataaaaggcg cctgtagtgc catttacccc cattcactgc
     7441 cagagccgtg agcgcagcga actgaatgtc acgaaaaaga cagcgactca ggtgcctgat
     7501 ggtcggagac aaaaggaata ttcagcgatt tgcccgagct tgcgagggtg ctacttaagc
     7561 ctttagggtt ttaaggtctg ttttgtagag gagcaaacag cgtttgcgac atccttttgt
     7621 aatactgcgg aactgactaa agtagtgagt tatacacagg gctgggatct attcttttta
     7681 tcttttttta ttctttcttt attctataaa ttataaccac ttgaatataa acaaaaaaaa
     7741 cacacaaagg tctagcggaa tttacagagg gtctagcaga atttacaagt tttccagcaa
     7801 aggtctagca gaatttacag atacccacaa ctcaaaggaa aaggactagt aattatcatt
     7861 gactagccca tctcaattgg tatagtgatt aaaattacct agaccaattg agatgtatgt
     7921 ctgaattagt tgttttcaaa gcaaatgaac tagcgattag tcgctatgac ttaacggagc
     7981 atgaaaccaa gctaatttta tgctgtgtgg cactactcaa ccccacgatt gaaaacccta
     8041 caaggaaaga acggacggta tcgttcactt ataaccaata cgctcagatg atgaacatca
     8101 gtagggaaaa tgcttatggt gtattagcta aagcaaccag agagctgatg acgagaactg
     8161 tgaaaatcag gaatcctttg gttaaaggct ttgagatttt ccagtggaca aactatgcca
     8221 agttctcaag cgaaaaatta gaattagttt ttagtgaaga gatattgcct tatcttttcc
     8281 agttaaaaaa attcataaaa tataatctgg aacatgttaa gtcttttgaa aacaaatact
     8341 ctatgaggat ttatgagtgg ttattaaaag aactaacaca aaagaaaact cacaaggcaa
     8401 atatagagat tagccttgat gaatttaagt tcatgttaat gcttgaaaat aactaccatg
     8461 agtttaaaag gcttaaccaa tgggttttga aaccaataag taaagattta aacacttaca
     8521 gcaatatgaa attggtggtt gataagcgag gccgcccgac tgatacgttg attttccaag
     8581 ttgaactaga tagacaaatg gatctcgtaa ccgaacttga gaacaaccag ataaaaatga
     8641 atggtgacaa aataccaaca accattacat cagattccta cctacgtaac ggactaagaa
     8701 aaacactaca cgatgcttta actgcaaaaa ttcagctcac cagttttgag gcaaaatttt
     8761 tgagtgacat gcaaagtaag catgatctca atggttcgtt ctcatggctc acgcaaaaac
     8821 aacgaaccac actagagaac atactggcta aatacggaag gatctga
//
