LOCUS       sequence-337586-        8918 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 PTP1B.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8918)
  TITLE     Fox lab plasmids
REFERENCE   2  (bases 1 to 8918)
  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: pB2Hopt
FEATURES             Location/Qualifiers
     source          1..8918
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..816
                     /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             1..816
                     /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(74..93)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     complement(862..879)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1032..1118
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1210..1237
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     CDS             2069..2092
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2412..2684
                     /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             4952..4963
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             complement(6007..6798)
                     /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      7487..7709
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             7757..8707
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVKIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
     primer_bind     complement(8728..8746)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        8814..8918
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atgagccata ttcaacggga aacgtcttgc tctaggccgc gattaaattc caacatggat
       61 gctgatttat atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc
      121 tatcgattgt atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc
      181 gttgccaatg atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct
      241 cttccgacca tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg
      301 atccccggga aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt
      361 gttgatgcgc tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct
      421 tttaacagcg atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg
      481 gttgatgcga gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa
      541 gaaatgcata aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca
      601 cttgataacc ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc
      661 ggaatcgcag accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct
      721 ccttcattac agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa
      781 ttgcagtttc atttgatgct cgatgagttt ttctaaacgc catgggcatg gctgttttgg
      841 cggatgagag aagattttca gcctgataca gattaaatca gaacgcagaa gcggtctgat
      901 aaaacagaat ttgcctggcg acagtagcgc ggtggtccca cctgacccca tgccgaactc
      961 agaagtgaaa cgccgtagcg ccgatggtag tgtggggtct ccccatgcga gagtagggaa
     1021 ctgccaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt cgttttatct
     1081 gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc gccgggagcg gatttgaacg
     1141 ttgcgaagca acggcccgga gggtggcggg caggacgccc gccataaact gccaggcatc
     1201 aaattaagca gaaggccatc ctgacggatg gcctttttgc gtttctacaa actcgctgag
     1261 gattctagag cacagctaac accacgtcgt ccctatctgc tgccctaggt ctatgagtgg
     1321 ttgctggata actttacggg catgcataag gctcggtatc tatattcagg gagaccacaa
     1381 cggtttccct ctacaaataa ttttgtttaa cttttactag agtaaggagg aaaaaaaaat
     1441 gagtatcagc agcagggtaa aaagcaaaag aattcagctt ggacttaacc aggctgaact
     1501 tgctcaaaag gtggggacta cccagcagtc tatagagcag ctcgaaaacg gtaaaactaa
     1561 gcgaccacgc tttttaccag aacttgcgtc agctcttggc gtaagtgttg actggctgct
     1621 caatggcacc tctgattcga atgttagatt tgttgggcac gttgagccca aagggaaata
     1681 tccattgatt agcatggtta gagctcgttc gtggtgtgaa gcttgtgaac cctacgatat
     1741 caaggacatt gatgaatggt atgacagtga cgttaactta ttaggcaatg gattctggct
     1801 gaaggttgaa ggtgattcca tgacctcacc tgtaggtcaa agcatccctg aaggtcatat
     1861 ggtgttagta gatactggac gggagccagt gaatggaagc cttgttgtag ccaaactgac
     1921 tgacgcgaac gaagcaacat tcaagaaact ggtcatagat ggcggtcaga agtacctgaa
     1981 aggcctgaat ccttcatggc ctatgactcc tatcaacgga aactgcaaga ttatcggtgt
     2041 tgtcgtggaa gcgagggtaa aattcgtaga ctacaaggac gacgatgaca agtggtattt
     2101 tgggaagatc actcgtcggg agtccgagcg gctgctgctc aaccccgaaa acccccgggg
     2161 aaccttcttg gtccgggaga gcgagacggt aaaaggtgcc tatgccctct ccgtttctga
     2221 ctttgacaac gccaaggggc tcaatgtgaa acactacctg atccgcaagc tggacagcgg
     2281 cggcttctac atcacctcac gcacacagtt cagcagcctg cagcagctgg tggcctacta
     2341 ctccaaacat gctgatggct tgtgccaccg cctgaccaac gtctgctaag tgcagtaagg
     2401 aggaaaaaaa aatggcacgc gtaactgttc aggacgctgt agagaaaatt ggtaaccgtt
     2461 ttgacctggt actggtcgcc gcgcgtcgcg ctcgtcagat gcaggtaggc ggaaaggatc
     2521 cgctggtacc ggaagaaaac gataaaacca ctgtaatcgc gctgcgcgaa atcgaagaag
     2581 gtctgatcaa caaccagatc ctcgacgttc gcgaacgcca ggaacagcaa gagcaggaag
     2641 ccgctgaatt acaagccgtt accgctattg ctgaaggtcg tcgtgcagct gcggaaccgc
     2701 agtatgaaga aattccgatt tatctgtaat aattctagag cacagctaac accacgtcgt
     2761 ccctatctgc tgccctaggt ctatgagtgg ttgctggata actttacggg catgcataag
     2821 gctcgtataa tatattcagg gagaccacaa cggtttccct ctacaaataa ttttgtttaa
     2881 cttttactag agcgaaaaaa agtaaggcgg taatccatgg gctccaagcc gcagactcag
     2941 ggcctggcca aggatgcctg ggagatccct cgggagtcgc tgcggctgga ggtcaagctg
     3001 ggccagggct gctttggcga ggtgtggatg gggacctgga acggtaccac cagggtggcc
     3061 atcaaaaccc tgaagcctgg cacgatgtct ccagaggcct tcctgcagga ggcccaggtc
     3121 atgaagaagc tgaggcatga gaagctggtg cagttgtatg ctgtggtttc agaggagccc
     3181 atttacatcg tcacggagta catgagcaag gggagtttgc tggactttct caagggggag
     3241 acaggcaagt acctgcggct gcctcagctg gtggacatgg ctgctcagat cgcctcaggc
     3301 atggcgtacg tggagcggat gaactacgtc caccgggacc ttcgtgcagc caacatcctg
     3361 gtgggagaga acctggtgtg caaagtggcc gactttgggc tggctcggct cattgaagac
     3421 aatgagtaca cggcgcggca aggtgccaaa ttccccatca agtggacggc tccagaagct
     3481 gccctctatg gccgcttcac catcaagtcg gacgtgtggt ccttcgggat cctgctgact
     3541 gagctcacca caaagggacg ggtgccctac cctgggatgg tgaaccgcga ggtgctggac
     3601 caggtggagc ggggctaccg gatgccctgc ccgccggagt gtcccgagtc cctgcacgac
     3661 ctcatgtgcc agtgctggcg gaaggagcct gaggagcggc ccaccttcga gtacctgcag
     3721 gccttcctgg aggactactt cacgtccacc gagccccagt accagcccgg ggagaacctc
     3781 taagctagct ttaagaagga gatataccat ggtggactac agcgtgtggg accacattga
     3841 ggtgtctgat gatgaagacg agacgcaccc caacatcgac acggccagtc tcttccgctg
     3901 gcggcatcag gcccgggtgg aacgcatgga gcagttccag aaggagaagg aggaactgga
     3961 caggggctgc cgcgagtgca agcgcaaggt ggccgagtgc cagaggaaac tgaaggagct
     4021 ggaggtggcc gagggcggca aggcagagct ggagcgcctg caggccgagg cacagcagct
     4081 gcgcaaggag gagcggagct gggagcagaa gctggaggag atgcgcaaga aggagaagag
     4141 catgccctgg aacgtggaca cgctcagcaa agacggcttc agcaagagca tggtaaatac
     4201 caagcccgag aagacggagg aggactcaga ggaggtgagg gagcagaaac acaagacctt
     4261 cgtggaaaaa tacgagaaac agatcaagca ctttggcatg cttcgccgct gggatgacag
     4321 ccaaaagtac ctgtcagaca acgtccacct ggtgtgcgag gagacagcca attacctggt
     4381 catttggtgc attgacctag aggtggagga gaaatgtgca ctcatggagc aggtggccca
     4441 ccagacaatc gtcatgcaat ttatcctgga gctggccaag agcctaaagg tggacccccg
     4501 ggcctgcttc cggcagttct tcactaagat taagacagcc gatcgccagt acatggaggg
     4561 cttcaacgac gagctggaag ccttcaagga gcgtgtgcgg ggccgtgcca agctgcgcat
     4621 cgagaaggcc atgaaggagt acgaggagga ggagcgcaag aagcggctcg gccccggcgg
     4681 cctggacccc gtcgaggtct acgagtccct ccctgaggaa ctccagaagt gcttcgatgt
     4741 gaaggacgtg cagatgctgc aggacgccat cagcaagatg gaccccaccg acgcaaagta
     4801 ccacatgcag cgctgcattg actctggcct ctgggtcccc aactctaagg ccagcgaggc
     4861 caaggaggga gaggaggcag gtcctgggga cccattactg gaagctgttc ccaagacggg
     4921 cgatgagaag gatgtcagtg tgtaagtgca gaaagaggag aaatactaga tggagatgga
     4981 aaaggagttc gagcagatcg acaagtccgg gagctgggcg gccatttacc aggatatccg
     5041 acatgaagcc agtgacttcc catgtagagt ggccaagctt cctaagaaca aaaaccgaaa
     5101 taggtacaga gacgtcagtc cctttgacca tagtcggatt aaactacatc aagaagataa
     5161 tgactatatc aacgctagtt tgataaaaat ggaagaagcc caaaggagtt acattcttac
     5221 ccagggccct ttgcctaaca catgcggtca cttttgggag atggtgtggg agcagaaaag
     5281 caggggtgtc gtcatgctca acagagtgat ggagaaaggt tcgttaaaat gcgcacaata
     5341 ctggccacaa aaagaagaaa aagagatgat ctttgaagac acaaatttga aattaacatt
     5401 gatctctgaa gatatcaagt catattatac agtgcgacag ctagaattgg aaaaccttac
     5461 aacccaagaa actcgcgaga tcttacattt ccactatacc acatggcctg actttggagt
     5521 ccctgaatca ccagcctcat tcttgaactt tcttttcaaa gtccgagagt cagggtcact
     5581 cagcccggag cacgggcccg ttgtggtgca ctgcagtgca ggcatcggca ggtctggaac
     5641 cttctgtctg gctgatacct gcctcttgct gatggacaag aggaaagacc cttcttccgt
     5701 tgatatcaag aaagtgctgt tagaaatgag gaagtttcgg atggggctga tccagacagc
     5761 cgaccagctg cgcttctcct acctggctgt gatcgaaggt gccaaattca tcatggggga
     5821 ctcttccgtg caggatcagt ggaaggagct ttcccacgag gacctggagc ccccacccga
     5881 gcatatcccc ccacctcccc ggccacccaa acgaatcctg gagccacaca attgacagtc
     5941 agtagggccc taaaaaaaaa ccccgcccct gacaggggtg gggttttttg cgtacattgg
     6001 ctccgttcat ttgccgacta ccttggtgat ctcgcctttc acgtagtgga caaattcttc
     6061 caactgatct gcgcgcgagg ccaagcgatc ttcttcttgt ccaagataag cctgtctagc
     6121 ttcaagtatg acgggctgat actgggccgg caggcgctcc attgcccagt cggcagcgac
     6181 atccttcggc gcgattttgc cggttactgc gctgtaccaa atgcgggaca acgtaagcac
     6241 tacatttcgc tcatcgccag cccagtcggg cggcgagttc catagcgtta aggtttcatt
     6301 tagcgcctca aatagatcct gttcaggaac cggatcaaag agttcctccg ccgctggacc
     6361 taccaaggca acgctatgtt ctcttgcttt tgtcagcaag atagccagat caatgtcgat
     6421 cgtggctggc tcgaagatac ctgcaagaat gtcattgcgc tgccattctc caaattgcag
     6481 ttcgcgctta gctggataac gccacggaat gatgtcgtcg tgcacaacaa tggtgacttc
     6541 tacagcgcgg agaatctcgc tctctccagg ggaagccgaa gtttccaaaa ggtcgttgat
     6601 caaagctcgc cgcgttgttt catcaagcct tacggtcacc gtaaccagca aatcaatatc
     6661 actgtgtggc ttcaggccgc catccactgc ggagccgtac aaatgtacgg ccagcaacgt
     6721 cggttcgaga tggcgctcga tgacgccaac tacctctgat agttgagtcg atacttcggc
     6781 gatcaccgct tccctcattt ttttttcctc cttactgcac ctgcaggtcg gtcgacacaa
     6841 catacgcgcc ggaagcttta cgagtaaagc ccggggtgcc taatgaacaa actttcttgt
     6901 attttcatac aatgtatctt gttttagggt ccctaagtct ccgcagcgca acgaaaggcc
     6961 cagtctttcg actgagcctt tcgttgcatt tgcccggatg tcctcttgac cgtacgacgg
     7021 gtaagcctgt tgatgatacc gctgccttac tgggtgcatt agccagtctg aatgacctgt
     7081 cacgggataa tccgaagtgg tcagactgga aaatcagagg gcaggaactg ctgaacagca
     7141 aaaagtcaga tagcaccaca tagcagaccc gccataaaac gccctgagaa gcccgtgacg
     7201 ggcttttctt gtattatggg tagtttcctt gcatgaatcc ataaaaggcg cctgtagtgc
     7261 catttacccc cattcactgc cagagccgtg agcgcagcga actgaatgtc acgaaaaaga
     7321 cagcgactca ggtgcctgat ggtcggagac aaaaggaata ttcagcgatt tgcccgagct
     7381 tgcgagggtg ctacttaagc ctttagggtt ttaaggtctg ttttgtagag gagcaaacag
     7441 cgtttgcgac atccttttgt aatactgcgg aactgactaa agtagtgagt tatacacagg
     7501 gctgggatct attcttttta tcttttttta ttctttcttt attctataaa ttataaccac
     7561 ttgaatataa acaaaaaaaa cacacaaagg tctagcggaa tttacagagg gtctagcaga
     7621 atttacaagt tttccagcaa aggtctagca gaatttacag atacccacaa ctcaaaggaa
     7681 aaggactagt aattatcatt gactagccca tctcaattgg tatagtgatt aaaattacct
     7741 agaccaattg agatgtatgt ctgaattagt tgttttcaaa gcaaatgaac tagcgattag
     7801 tcgctatgac ttaacggagc atgaaaccaa gctaatttta tgctgtgtgg cactactcaa
     7861 ccccacgatt gaaaacccta caaggaaaga acggacggta tcgttcactt ataaccaata
     7921 cgctcagatg atgaacatca gtagggaaaa tgcttatggt gtattagcta aagcaaccag
     7981 agagctgatg acgagaactg tgaaaatcag gaatcctttg gttaaaggct ttgagatttt
     8041 ccagtggaca aactatgcca agttctcaag cgaaaaatta gaattagttt ttagtgaaga
     8101 gatattgcct tatcttttcc agttaaaaaa attcataaaa tataatctgg aacatgttaa
     8161 gtcttttgaa aacaaatact ctatgaggat ttatgagtgg ttattaaaag aactaacaca
     8221 aaagaaaact cacaaggcaa atatagagat tagccttgat gaatttaagt tcatgttaat
     8281 gcttgaaaat aactaccatg agtttaaaag gcttaaccaa tgggttttga aaccaataag
     8341 taaagattta aacacttaca gcaatatgaa attggtggtt gataagcgag gccgcccgac
     8401 tgatacgttg attttccaag ttgaactaga tagacaaatg gatctcgtaa ccgaacttga
     8461 gaacaaccag ataaaaatga atggtgacaa aataccaaca accattacat cagattccta
     8521 cctacgtaac ggactaagaa aaacactaca cgatgcttta actgcaaaaa ttcagctcac
     8581 cagttttgag gcaaaatttt tgagtgacat gcaaagtaag catgatctca atggttcgtt
     8641 ctcatggctc acgcaaaaac aacgaaccac actagagaac atactggcta aatacggaag
     8701 gatctgaacg gcctatttgg cctaagggcc tcgtgatacg cctattttta taggttaatg
     8761 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     8821 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     8881 cctgataaat gcttcaataa tattgaaaaa ggaagagt
//
