LOCUS       sequence-361438-        5727 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5727)
  AUTHORS   Emrick MA, Hoofnagle AN, Miller AS, Ten Eyck LF, Ahn NG
  TITLE     Constitutive activation of extracellular signal-regulated kinase 2 
            by synergistic point mutations.
  JOURNAL   J Biol Chem. 2001 Dec 7;276(49):46469-79. Epub 2001 Oct 8.
  PUBMED    11591711
REFERENCE   2  (bases 1 to 5727)
  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: "J Biol 
            Chem."; date: "2001-12-7"; volume: "276(49)"; pages: "46469-79. Epub
            2001 Oct 8"
COMMENT     Sequence Label: pCMV5-rat ERK2-L73P/S151D
FEATURES             Location/Qualifiers
     source          1..5727
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     RBS             608..616
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             629..646
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     regulatory      648..657
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1685..1720
                     /codon_start=1
                     /product="epitope tag derived from MAP kinase 2"
                     /label=Tag-100
                     /translation="EETARFQPGYRS"
     polyA_signal    1751..2373
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(1950..1969)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2397..2417)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2402..2731
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2582..2717
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2769..2788)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2770..2788)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2802..2818)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2802..2818)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2815..2837)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2826..2842
                     /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)."
     promoter        complement(join(2850..2856,2857..2874,2875..2880))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2895..2916
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3033..3050)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3204..3792)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3284..3303)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3963..4754,4755..4823))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4586..4605
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4824..4928)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4955..5410
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5042..5061)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5252..5273
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5536..5558
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5550..5567
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5551..5567
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agaattctaa ggaggtttaa ccatggcaca tcaccatcac catcacgcca tggcggcggc
      661 ggcggcgggc ccggagatgg tccgcgggca ggtgttcgac gtggggccgc gctacactaa
      721 tctctcgtac atcggagaag gcgcctacgg catggtttgt tctgcttatg ataatctcaa
      781 caaagttcga gttgctatca agaaaatcag tccttttgag caccagacct actgtcagag
      841 aaccctgaga gagataaaaa tcccactgcg cttcagacat gagaatatta tcggcatcaa
      901 tgacatcatc cgggcaccaa ccattgagca gatgaaagat gtatatatag tacaggacct
      961 catggagaca gatctttaca agctcttgaa gacacagcac ctcagcaatg atcatatctg
     1021 ctattttctt tatcagatcc tgagaggatt aaagtatata cattcagcta atgttctgca
     1081 ccgtgacctt aagcctgaca acctcctgct gaacaccact tgtgatctca agatctgtga
     1141 ctttggcctt gcccgtgttg cagatccaga ccatgatcat acagggttct tgacagagta
     1201 tgtagccacg cgttggtaca gagctccaga aattatgttg aattccaagg gttataccaa
     1261 gtccattgat atttggtctg tgggctgcat cctggcagag atgctatcca acaggcctat
     1321 cttcccagga aagcattacc ttgaccagct gaatcacatc ctgggtattc ttggatctcc
     1381 atcacaggaa gatctgaatt gtataataaa tttaaaagct agaaactatt tgctttctct
     1441 cccgcacaaa aataaggtgc cgtggaacag attgttccca aacgctgact ccaaagctct
     1501 ggatttactg gataaaatgt tgacatttaa ccctcacaag aggattgaag ttgaacaggc
     1561 tctggcccac ccgtacctgg agcagtatta tgacccaagt gatgagccca ttgctgaagc
     1621 accattcaag tttgacatgg agctggacga cttacctaag gagaagctca aagaactcat
     1681 ttttgaagag actgctcgat tccagccagg atacagatct taacaagcgg ccgcgactct
     1741 agaggatccc gggtggcatc cctgtgaccc ctccccagtg cctctcctgg ccctggaagt
     1801 tgccactcca gtgcccacca gccttgtcct aataaaatta agttgcatca ttttgtctga
     1861 ctaggtgtcc ttctataata ttatggggtg gaggggggtg gtatggagca aggggcaagt
     1921 tgggaagaca acctgtaggg cctgcggggt ctattgggaa ccaagctgga gtgcagtggc
     1981 acaatcttgg ctcactgcaa tctccgcctc ctgggttcaa gcgattctcc tgcctcagcc
     2041 tcccgagttg ttgggattcc aggcatgcat gaccaggctc agctaatttt tgtttttttg
     2101 gtagagacgg ggtttcacca tattggccag gctggtctcc aactcctaat ctcaggtgat
     2161 ctacccacct tggcctccca aattgctggg attacaggcg tgaaccactg ctcccttccc
     2221 tgtccttctg attttaaaat aactatacca gcaggaggac gtccagacac agcataggct
     2281 acctggccat gcccaaccgg tgggacattt gagttgcttg cttggcactg tcctctcatg
     2341 cgttgggtcc actcagtaga tgcctgttga attgggtacg cggccagctt ggctgtggaa
     2401 tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     2461 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     2521 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     2581 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     2641 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     2701 aggctttttt ggaggcctag gcttttgcaa aaagctcctc gaggaactga aaaaccagaa
     2761 agttaattcc ctatagtgag tcgtattaaa ttcgtaatca tgtcatagct gtttcctgtg
     2821 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     2881 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     2941 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     3001 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     3061 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     3121 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     3181 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     3241 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     3301 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     3361 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     3421 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     3481 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     3541 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     3601 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     3661 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     3721 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     3781 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     3841 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     3901 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     3961 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     4021 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     4081 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     4141 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     4201 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     4261 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     4321 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     4381 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     4441 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     4501 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     4561 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     4621 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     4681 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     4741 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     4801 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     4861 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     4921 gttccgcgca catttccccg aaaagtgcca cctgacgcgc cctgtagcgg cgcattaagc
     4981 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     5041 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     5101 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     5161 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     5221 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     5281 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     5341 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     5401 cttacaattt ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     5461 cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg
     5521 taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagctgatct
     5581 atacattgaa tcaatattgg caattagcca tattagtcat tggttatata gcataaatca
     5641 atattggcta ttggccattg catacgttgt atctatatca taatatgtac atttatattg
     5701 gctcatgtcc aatatgaccg ccatgtt
//
