LOCUS       sequence-337609-        8873 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 PTPN12 (Y64A).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8873)
  TITLE     Fox lab plasmids
REFERENCE   2  (bases 1 to 8873)
  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: pB2H12*
FEATURES             Location/Qualifiers
     source          1..8873
                     /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(6173..6964)
                     /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      7653..7875
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             7923..8873
                     /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 atggagcaag tggagatcct gaggaaattc atccagaggg
     5221 tccaggccat gaagagtcct gaccacaatg gggaggacaa cttcgcccgg gacttcatgc
     5281 ggttaagaag attgtctacc aaatatagaa cagaaaagat atatcccaca gccactggag
     5341 aaaaagaaga aaatgttaaa aagaacagag cgaaggacat actgccattt gatcacagcc
     5401 gagttaaatt gacattaaag actccttcac aagattcaga ctatatcaat gcaaatttta
     5461 taaagggcgt ctatgggcca aaagcatatg tagcaactca aggaccttta gcaaatacag
     5521 taatagattt ttggaggatg atatgggagt ataatgttgt gatcattgta atggcctgcc
     5581 gagaatttga gatgggaagg aaaaaatgtg agcgctattg gcctttgtat ggagaagacc
     5641 ccataacgtt tgcaccattt aaaatttctt gtgaggatga acaagcaaga acagactact
     5701 tcatcaggac actcttactt gaatttcaaa atgaatctcg taggctgtat cagtttcatt
     5761 atgtgaactg gccagaccat gatgttcctt catcatttga ttctattctg gacatgataa
     5821 gcttaatgag gaaatatcaa gaacatgaag atgttcctat ttgtattcat tgcagtgcag
     5881 gctgtggaag aacaggtgcc atttgtgcca tagattatac gtggaattta ctaaaagctg
     5941 ggaaaatacc agaggaattt aatgtattta atttaataca agaaatgaga acacaaaggc
     6001 attctgcagt acaaacaaag gagcaatatg aacttgttca tagagctatt gcccaactgt
     6061 ttgaaaaaca gctacaacta tatgaaattc atggagctta acagtcagta gggccctaaa
     6121 aaaaaacccc gcccctgaca ggggtggggt tttttgcgta cattggctcc gttcatttgc
     6181 cgactacctt ggtgatctcg cctttcacgt agtggacaaa ttcttccaac tgatctgcgc
     6241 gcgaggccaa gcgatcttct tcttgtccaa gataagcctg tctagcttca agtatgacgg
     6301 gctgatactg ggccggcagg cgctccattg cccagtcggc agcgacatcc ttcggcgcga
     6361 ttttgccggt tactgcgctg taccaaatgc gggacaacgt aagcactaca tttcgctcat
     6421 cgccagccca gtcgggcggc gagttccata gcgttaaggt ttcatttagc gcctcaaata
     6481 gatcctgttc aggaaccgga tcaaagagtt cctccgccgc tggacctacc aaggcaacgc
     6541 tatgttctct tgcttttgtc agcaagatag ccagatcaat gtcgatcgtg gctggctcga
     6601 agatacctgc aagaatgtca ttgcgctgcc attctccaaa ttgcagttcg cgcttagctg
     6661 gataacgcca cggaatgatg tcgtcgtgca caacaatggt gacttctaca gcgcggagaa
     6721 tctcgctctc tccaggggaa gccgaagttt ccaaaaggtc gttgatcaaa gctcgccgcg
     6781 ttgtttcatc aagccttacg gtcaccgtaa ccagcaaatc aatatcactg tgtggcttca
     6841 ggccgccatc cactgcggag ccgtacaaat gtacggccag caacgtcggt tcgagatggc
     6901 gctcgatgac gccaactacc tctgatagtt gagtcgatac ttcggcgatc accgcttccc
     6961 tcattttttt ttcctcctta ctgcacctgc aggtcggtcg acacaacata cgcgccggaa
     7021 gctttacgag taaagcccgg ggtgcctaat gaacaaactt tcttgtattt tcatacaatg
     7081 tatcttgttt tagggtccct aagtctccgc agcgcaacga aaggcccagt ctttcgactg
     7141 agcctttcgt tgcatttgcc cggatgtcct cttgaccgta cgacgggtaa gcctgttgat
     7201 gataccgctg ccttactggg tgcattagcc agtctgaatg acctgtcacg ggataatccg
     7261 aagtggtcag actggaaaat cagagggcag gaactgctga acagcaaaaa gtcagatagc
     7321 accacatagc agacccgcca taaaacgccc tgagaagccc gtgacgggct tttcttgtat
     7381 tatgggtagt ttccttgcat gaatccataa aaggcgcctg tagtgccatt tacccccatt
     7441 cactgccaga gccgtgagcg cagcgaactg aatgtcacga aaaagacagc gactcaggtg
     7501 cctgatggtc ggagacaaaa ggaatattca gcgatttgcc cgagcttgcg agggtgctac
     7561 ttaagccttt agggttttaa ggtctgtttt gtagaggagc aaacagcgtt tgcgacatcc
     7621 ttttgtaata ctgcggaact gactaaagta gtgagttata cacagggctg ggatctattc
     7681 tttttatctt tttttattct ttctttattc tataaattat aaccacttga atataaacaa
     7741 aaaaaacaca caaaggtcta gcggaattta cagagggtct agcagaattt acaagttttc
     7801 cagcaaaggt ctagcagaat ttacagatac ccacaactca aaggaaaagg actagtaatt
     7861 atcattgact agcccatctc aattggtata gtgattaaaa ttacctagac caattgagat
     7921 gtatgtctga attagttgtt ttcaaagcaa atgaactagc gattagtcgc tatgacttaa
     7981 cggagcatga aaccaagcta attttatgct gtgtggcact actcaacccc acgattgaaa
     8041 accctacaag gaaagaacgg acggtatcgt tcacttataa ccaatacgct cagatgatga
     8101 acatcagtag ggaaaatgct tatggtgtat tagctaaagc aaccagagag ctgatgacga
     8161 gaactgtgaa aatcaggaat cctttggtta aaggctttga gattttccag tggacaaact
     8221 atgccaagtt ctcaagcgaa aaattagaat tagtttttag tgaagagata ttgccttatc
     8281 ttttccagtt aaaaaaattc ataaaatata atctggaaca tgttaagtct tttgaaaaca
     8341 aatactctat gaggatttat gagtggttat taaaagaact aacacaaaag aaaactcaca
     8401 aggcaaatat agagattagc cttgatgaat ttaagttcat gttaatgctt gaaaataact
     8461 accatgagtt taaaaggctt aaccaatggg ttttgaaacc aataagtaaa gatttaaaca
     8521 cttacagcaa tatgaaattg gtggttgata agcgaggccg cccgactgat acgttgattt
     8581 tccaagttga actagataga caaatggatc tcgtaaccga acttgagaac aaccagataa
     8641 aaatgaatgg tgacaaaata ccaacaacca ttacatcaga ttcctaccta cgtaacggac
     8701 taagaaaaac actacacgat gctttaactg caaaaattca gctcaccagt tttgaggcaa
     8761 aatttttgag tgacatgcaa agtaagcatg atctcaatgg ttcgttctca tggctcacgc
     8821 aaaaacaacg aaccacacta gagaacatac tggctaaata cggaaggatc tga
//
