LOCUS       sequence-460691-        6357 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST-HA tagged human MEK5 from bacterial expression vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6357)
  AUTHORS   Brady DC, Crowe MS, Turski ML, Hobbs GA, Yao X, Chaikuad A, Knapp S,
            Xiao K, Campbell SL, Thiele DJ, Counter CM
  TITLE     Copper is required for oncogenic BRAF signalling and tumorigenesis.
  JOURNAL   Nature. 2014 Apr 9. doi: 10.1038/nature13180.
  PUBMED    24717435
REFERENCE   2  (bases 1 to 6357)
  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: "Nature. 
            2014 Apr 9. doi: 10.1038/nature13180."
COMMENT     Sequence Label: pGEX6P1-HA-MEK5
FEATURES             Location/Qualifiers
     source          1..6357
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..22,23..29)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    37..53
                     /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             76..729
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     687..709
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             736..759
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             781..807
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2225..2247)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2392..2410)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2478..2582
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2583..2651,2652..3443)
                     /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(2801..2820)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3614..4202
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4103..4122
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4356..4373
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4446..4523
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4524..5606
                     /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     complement(4543..4562)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5655..5660,5661..5678,5679..5685)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5693..5709
                     /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     5717..5733
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5717..5733
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             5729..6256
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5749..5766)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5749..5765)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5758..5780)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca
       61 caggaaacag tattcatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa
      121 cccactcgac ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc
      181 gatgaaggtg ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt
      241 ccttattata ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata
      301 gctgacaagc acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt
      361 gaaggagcgg ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt
      421 gaaactctca aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat
      481 cgtttatgtc ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg
      541 tatgacgctc ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa
      601 ttagtttgtt ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc
      661 agcaagtata tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat
      721 cctccaaaat cggatctgga agttctgttc caggggcccc tgggatccgc cgccaccatg
      781 tacccatacg atgttccaga ttacgctctg tggctagccc ttggcccctt tcctgccatg
      841 gagaaccagg tgctggtaat tcgcatcaag atcccaaata gtggcgcggt ggactggaca
      901 gtgcactccg ggccgcagtt actcttcagg gatgtgctgg atgtgatagg ccaggttctg
      961 cctgaagcaa caactacagc atttgaatat gaagatgaag atggtgatcg aattacagtg
     1021 agaagtgatg aggaaatgaa ggcaatgctg tcatattatt attccacagt aatggaacag
     1081 caagtaaatg gacagttaat agagcctctg cagatatttc caagagcctg caagcctcct
     1141 ggggaacgga acatacatgg cctgaaggtg aatactcggg ccggaccctc tcaacacagc
     1201 agcccagcag tctcagattc acttccaagc aatagcttaa agaagtcttc tgctgaactg
     1261 aaaaaaatac tagccaatgg ccagatgaat gaacaagaca tacgatatcg ggacactctt
     1321 ggtcatggca acggaggcac agtctacaaa gcatatcatg tcccgagtgg gaaaatatta
     1381 gctgtaaagg tcatactact agatattaca ctggaacttc agaagcaaat tatgtctgaa
     1441 ttggaaattc tttataagtg cgattcatca tatatcattg gattttatgg agcatttttt
     1501 gtagaaaaca ggatttcaat atgtacagaa ttcatggatg ggggatcttt ggatgtatat
     1561 aggaaaatgc cagaacatgt ccttggaaga attgcagtag cagttgttaa aggccttact
     1621 tatttgtgga gtttaaagat tttacataga gacgtgaagc cctccaatat gctagtaaac
     1681 acaagaggac aggttaagct gtgtgatttt ggagttagca ctcagctggt gaattctata
     1741 gccaagacgt atgttggaac aaatgcttat atggcgcctg aaaggatttc aggggagcag
     1801 tatggaattc attctgatgt ctggagctta ggaatctctt ttatggagct tgctcttggg
     1861 aggtttccat atcctcagat tcagaaaaac cagggatctt taatgcctct ccagcttctg
     1921 cagtgcattg ttgatgagga ttcgcccgtc cttccagttg gagagttctc ggagccattt
     1981 gtacatttca tcactcagtg tatgcgaaaa cagccaaaag aaaggccagc acctgaagaa
     2041 ttgatgggcc acccgttcat cgtgcagttc aatgatggaa atgccgccgt ggtgtccatg
     2101 tgggtgtgcc gggcgctgga ggagaggcgg agccagcagc aggggccccc gtgagtcgac
     2161 tcgagcggcc gcatcgtgac tgactgacga tctgcctcgc gcgtttcggt gatgacggtg
     2221 aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg
     2281 ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca
     2341 tgacccagtc acgtagcgat agcggagtgt ataattcttg aagacgaaag ggcctcgtga
     2401 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     2461 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     2521 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     2581 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     2641 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     2701 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     2761 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     2821 cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     2881 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     2941 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     3001 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     3061 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     3121 tgcctgcagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     3181 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     3241 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     3301 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     3361 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     3421 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     3481 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     3541 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     3601 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     3661 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     3721 aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt
     3781 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     3841 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     3901 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     3961 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     4021 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     4081 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     4141 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     4201 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     4261 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
     4321 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     4381 aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcataa
     4441 attccgacac catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag cgcccggaag
     4501 agagtcaatt cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc gcagagtatg
     4561 ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac gtttctgcga
     4621 aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg
     4681 cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc agtctggccc
     4741 tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca
     4801 gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca
     4861 atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat gaccaggatg
     4921 ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat gtctctgacc
     4981 agacacccat caacagtatt attttctccc atgaagacgg tacgcgactg ggcgtggagc
     5041 atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggcccatta agttctgtct
     5101 cggcgcgtct gcgtctggct ggctggcata aatatctcac tcgcaatcaa attcagccga
     5161 tagcggaacg ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc
     5221 tgaatgaggg catcgttccc actgcgatgc tggttgccaa cgatcagatg gcgctgggcg
     5281 caatgcgcgc cattaccgag tccgggctgc gcgttggtgc ggatatctcg gtagtgggat
     5341 acgacgatac cgaagacagc tcatgttata tcccgccgtt aaccaccatc aaacaggatt
     5401 ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca actctctcag ggccaggcgg
     5461 tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag aaaaaccacg ctagcgccca
     5521 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
     5581 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtaagtta gctcactcat
     5641 taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc
     5701 ggataacaat ttcacacagg aaacagctat gaccatgatt acggattcac tggccgtcgt
     5761 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     5821 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     5881 gttgcgcagc ctgaatggcg aatggcgctt tgcctggttt ccggcaccag aagcggtgcc
     5941 ggaaagctgg ctggagtgcg atcttcctga ggccgatact gtcgtcgtcc cctcaaactg
     6001 gcagatgcac ggttacgatg cgcccatcta caccaacgtg acctatccca ttacggtcaa
     6061 tccgccgttt gttcccacgg agaatccgac gggttgttac tcgctcacat ttaatgttga
     6121 tgaaagctgg ctacaggaag gccagacgcg aattattttt gatggcgttg gaattagctt
     6181 atcgactgca cggtgcacca atgcttctgg cgtcaggcag ccatcggaag ctgtggtatg
     6241 gctgtgcagg tcgtaaatca ctgcataatt cgtgtcgctc aaggcgcact cccgttctgg
     6301 ataatgtttt ttgcgccgac atcataacgg ttctggcaaa tattctgaaa tgagctg
//
