LOCUS       sequence-310546-        8265 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Luminescent reporter of ERK activity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8265)
  AUTHORS   Day EK, Sosale NG, Xiao A, Zhong Q, Purow B, Lazzara MJ
  TITLE     Glioblastoma Cell Resistance to EGFR and MET Inhibition Can Be 
            Overcome via Blockade of FGFR-SPRY2 Bypass Signaling.
  JOURNAL   Cell Rep. 2020 Mar 10;30(10):3383-3396.e7. doi: 
            10.1016/j.celrep.2020.02.014.
  PUBMED    32160544
REFERENCE   2  (bases 1 to 8265)
  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"; doi: 
            "10.1016/j.celrep.2020.02"; journalName: "Cell Rep"; date: 
            "2020-03-10- 10"; volume: "30"; issue: "10"; pages: "3383-3396.e7"
COMMENT     Sequence Label: pMSCV-NLS-CBGreen-FIRE-puro
FEATURES             Location/Qualifiers
     source          1..8265
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..517
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    581..922
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             989..1405
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1335..1357
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1373..1389
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1418..1427
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1430..1450
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1451..3076
                     /codon_start=1
                     /product="green-emitting variant of click beetle 
                     luciferase"
                     /label=CBG99luc
                     /note="human codon-optimized"
                     /translation="MVKREKNVIYGPEPLHPLEDLTAGEMLFRALRKHSHLPQALVDVV
                     GDESLSYKEFFEATVLLAQSLHNCGYKMNDVVSICAENNTRFFIPVIAAWYIGMIVAPV
                     NESYIPDELCKVMGISKPQIVFTTKNILNKVLEVQSRTNFIKRIIILDTVENIHGCESL
                     PNFISRYSDGNIANFKPLHFDPVEQVAAILCSSGTTGLPKGVMQTHQNICVRLIHALDP
                     RVGTQLIPGVTVLVYLPFFHAFGFSITLGYFMVGLRVIMFRRFDQEAFLKAIQDYEVRS
                     VINVPSVILFLSKSPLVDKYDLSSLRELCCGAAPLAKEVAEVAAKRLNLPGIRCGFGLT
                     ESTSANIHSLGDEFKSGSLGRVTPLMAAKIADRETGKALGPNQVGELCIKGPMVSKGYV
                     NNVEATKEAIDDDGWLHSGDFGYYDEDEHFYVVDRYKELIKYKGSQVAPAELEEILLKN
                     PCIRDVAVVGIPDLEAGELPSAFVVKQPGKEITAKEVYDYLAERVSHTKYLRGGVRFVD
                     SIPRNVTGKITRKELLKQLLEKAGG"
     RBS             2401..2409
                     /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)"
     promoter        3441..3940
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3476..3499)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3873..3892
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3961..4560
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3961..3980)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4457..4477
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4660..5174
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5336..5352
                     /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(5360..5366,5367..5384,5385..5390))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5405..5426
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5543..5560)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5714..6302)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5794..5813)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6473..7264,7265..7333))
                     /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     7096..7115
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7334..7438)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7506..7524
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7562..7584)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7684..7703
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7897..7919
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8047..8066
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc
       61 atggaaaata cataactgag aatagagaag ttcagatcaa ggttaggaac agagagacag
      121 cagaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa
      181 gaacagatgg tccccagatg cggtcccgcc ctcagcagtt tctagagaac catcagatgt
      241 ttccagggtg ccccaaggac ctgaaatgac cctgtgcctt atttgaacta accaatcagt
      301 tcgcttctcg cttctgttcg cgcgcttctg ctccccgagc tcaataaaag agcccacaac
      361 ccctcactcg gcgcgccagt cctccgatag actgcgtcgc ccgggtaccc gtattcccaa
      421 taaagcctct tgctgtttgc atccgaatcg tggactcgct gatccttggg agggtctcct
      481 cagattgatt gactgcccac ctcgggggtc tttcatttgg aggttccacc gagatttgga
      541 gacccctgcc cagggaccac cgaccccccc gccgggaggt aagctggcca gcggtcgttt
      601 cgtgtctgtc tctgtctttg tgcgtgtttg tgccggcatc taatgtttgc gcctgcgtct
      661 gtactagtta gctaactagc tctgtatctg gcggacccgt ggtggaactg acgagttctg
      721 aacacccggc cgcaaccctg ggagacgtcc cagggacttt gggggccgtt tttgtggccc
      781 gacctgagga agggagtcga tgtggaatcc gaccccgtca ggatatgtgg ttctggtagg
      841 agacgagaac ctaaaacagt tcccgcctcc gtctgaattt ttgctttcgg tttggaaccg
      901 aagccgcgcg tcttgtctgc tgcagcgctg cagcatcgtt ctgtgttgtc tctgtctgac
      961 tgtgtttctg tatttgtctg aaaattaggg ccagactgtt accactccct taagtttgac
     1021 cttaggtcac tggaaagatg tcgagcggat cgctcacaac cagtcggtag atgtcaagaa
     1081 gagacgttgg gttaccttct gctctgcaga atggccaacc tttaacgtcg gatggccgcg
     1141 agacggcacc tttaaccgag acctcatcac ccaggttaag atcaaggtct tttcacctgg
     1201 cccgcatgga cacccagacc aggtccccta catcgtgacc tgggaagcct tggcttttga
     1261 cccccctccc tgggtcaagc cctttgtaca ccctaagcct ccgcctcctc ttcctccatc
     1321 cgccccgtct ctcccccttg aacctcctcg ttcgaccccg cctcgatcct ccctttatcc
     1381 agccctcact ccttctctag gcgccggaat tagatctgcc accatggccc ccaagaagaa
     1441 gaggaaggtg atggtgaagc gtgagaaaaa tgtcatctat ggccctgagc ctctccatcc
     1501 tttggaggat ttgactgccg gcgaaatgct gtttcgtgct ctccgcaagc actctcattt
     1561 gcctcaagcc ttggtcgatg tggtcggcga tgaatctttg agctacaagg agttttttga
     1621 ggcaaccgtc ttgctggctc agtccctcca caattgtggc tacaagatga acgacgtcgt
     1681 tagtatctgt gctgaaaaca atacccgttt cttcattcca gtcatcgccg catggtatat
     1741 cggtatgatc gtggctccag tcaacgagag ctacattccc gacgaactgt gtaaagtcat
     1801 gggtatctct aagccacaga ttgtcttcac cactaagaat attctgaaca aagtcctgga
     1861 agtccaaagc cgcaccaact ttattaagcg tatcatcatc ttggacactg tggagaatat
     1921 tcacggttgc gaatctttgc ctaatttcat ctctcgctat tcagacggca acatcgcaaa
     1981 ctttaaacca ctccacttcg accctgtgga acaagttgca gccattctgt gtagcagcgg
     2041 tactactgga ctcccaaagg gagtcatgca gacccatcaa aacatttgcg tgcgtctgat
     2101 ccatgctctc gatccacgcg tgggcactca gctgattcct ggtgtcaccg tcttggtcta
     2161 cttgcctttc ttccatgctt tcggctttag cattactttg ggttacttta tggtcggtct
     2221 ccgcgtgatt atgttccgcc gttttgatca ggaggctttc ttgaaagcca tccaagatta
     2281 tgaagtccgc agtgtcatca acgtgcctag cgtgatcctg tttttgtcta agagcccact
     2341 cgtggacaag tacgacttgt cttcactgcg tgaattgtgt tgcggtgccg ctccactggc
     2401 taaggaggtc gctgaagtgg ccgccaaacg cttgaatctt ccagggattc gttgtggctt
     2461 cggcctcacc gaatctacca gcgctaacat tcactctctc ggggatgagt ttaagagcgg
     2521 ctctttgggc cgtgtcactc cactcatggc tgctaagatc gctgatcgcg aaactggtaa
     2581 ggctttgggc ccgaaccaag tgggcgagct gtgtatcaaa ggccctatgg tgagcaaggg
     2641 ttatgtcaat aacgttgaag ctaccaagga ggccatcgac gacgacggct ggttgcattc
     2701 tggtgatttt ggatattacg acgaagatga gcatttttac gtcgtggatc gttacaagga
     2761 gctgatcaaa tacaagggta gccaggttgc tccagctgag ttggaggaga ttctgttgaa
     2821 aaatccatgc attcgcgatg tcgctgtggt cggcattcct gatctggagg ccggcgaact
     2881 gccttctgct ttcgttgtca agcagcctgg taaagaaatt accgccaaag aagtgtatga
     2941 ttacctggct gaacgtgtga gccatactaa gtacttgcgt ggcggcgtgc gttttgttga
     3001 ctccatccct cgtaacgtaa caggcaaaat tacccgcaag gagctgttga aacaattgtt
     3061 ggagaaggcc ggcggtttgt acaaaggcag cggcggatcc ctggtgctgg aagcccaccg
     3121 acccatctgc aaaatcccgg aaggagccaa ggagggggac acaggcagta ccagtggcac
     3181 cagcagccca ccagccccct gccgccctgt accttgtatc tccctttccc cagggcctgt
     3241 gcttgaacct gaggcactgc acacccccac actcatgacc acaccctccc taactccttt
     3301 cacccccagc ctggtcttca cctaccccag cactcctgag ccttgtgcct cagctcatcg
     3361 caagagtagc agcagcagcg gagacccatc ctctgacccc cttggctctc caaccctcct
     3421 cgctttgtga gaattctacc gggtagggga ggcgcttttc ccaaggcagt ctggagcatg
     3481 cgctttagca gccccgctgg gcacttggcg ctacacaagt ggcctctggc ctcgcacaca
     3541 ttccacatcc accggtaggc gccaaccggc tccgttcttt ggtggcccct tcgcgccacc
     3601 ttctactcct cccctagtca ggaagttccc ccccgccccg cagctcgcgt cgtgcaggac
     3661 gtgacaaatg gaagtagcac gtctcactag tctcgtgcag atggacagca ccgctgagca
     3721 atggaagcgg gtaggccttt ggggcagcgg ccaatagcag ctttgctcct tcgctttctg
     3781 ggctcagagg ctgggaaggg gtgggtccgg gggcgggctc aggggcgggc tcaggggcgg
     3841 ggcgggcgcc cgaaggtcct ccggaggccc ggcattctgc acgcttcaaa agcgcacgtc
     3901 tgccgcgctg ttctcctctt cctcatctcc gggcctttcg acctgcagcc caagcttacc
     3961 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     4021 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     4081 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     4141 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     4201 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     4261 tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag
     4321 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     4381 agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg
     4441 gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     4501 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga
     4561 cgcccgcccc acgacccgca gcgcccgacc gaaaggagcg cacgacccca tgcatcgata
     4621 aaataaaaga ttttatttag tctccagaaa aaggggggaa tgaaagaccc cacctgtagg
     4681 tttggcaagc tagcttaagt aacgccattt tgcaaggcat ggaaaataca taactgagaa
     4741 tagagaagtt cagatcaagg ttaggaacag agagacagca gaatatgggc caaacaggat
     4801 atctgtggta agcagttcct gccccggctc agggccaaga acagatggtc cccagatgcg
     4861 gtcccgccct cagcagtttc tagagaacca tcagatgttt ccagggtgcc ccaaggacct
     4921 gaaatgaccc tgtgccttat ttgaactaac caatcagttc gcttctcgct tctgttcgcg
     4981 cgcttctgct ccccgagctc aataaaagag cccacaaccc ctcactcggc gcgccagtcc
     5041 tccgatagac tgcgtcgccc gggtacccgt gtatccaata aaccctcttg cagttgcatc
     5101 cgacttgtgg tctcgctgtt ccttgggagg gtctcctctg agtgattgac tacccgtcag
     5161 cgggggtctt tcatgggtaa cagtttcttg aagttggaga acaacattct gagggtagga
     5221 gtcgaatatt aagtaatcct gactcaatta gccactgttt tgaatccaca tactccaata
     5281 ctcctgaaat agttcattat ggacagcgca gaaagagctg gggagaattg tgaaattgtt
     5341 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     5401 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     5461 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     5521 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     5581 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     5641 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     5701 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     5761 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     5821 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     5881 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     5941 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     6001 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     6061 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     6121 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     6181 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     6241 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     6301 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     6361 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     6421 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     6481 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     6541 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     6601 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     6661 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     6721 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     6781 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     6841 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     6901 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     6961 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     7021 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     7081 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     7141 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     7201 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     7261 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     7321 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     7381 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     7441 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     7501 ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa
     7561 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     7621 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact
     7681 atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca
     7741 gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc attcaggctg cgcaactgtt
     7801 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
     7861 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
     7921 cggcgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg ccacggggcc
     7981 tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag cccgatcttc
     8041 cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc
     8101 ggccacgatg cgtccggcgt agaggcgatt agtccaattt gttaaagaca ggatatcagt
     8161 ggtccaggct ctagttttga ctcaacaata tcaccagctg aagcctatag agtacgagcc
     8221 atagataaaa taaaagattt tatttagtct ccagaaaaag ggggg
//
