LOCUS       sequence-341824-        8678 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses MDA5 with a mutation to motif I in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8678)
  AUTHORS   Bamming D, Horvath CM
  TITLE     Regulation of signal transduction by enzymatically inactive 
            antiviral RNA helicase proteins MDA5, RIG-I, and LGP2.
  JOURNAL   J Biol Chem. 2009 Apr 10;284(15):9700-12. doi: 
            10.1074/jbc.M807365200. Epub 2009 Feb 11.
  PUBMED    19211564
REFERENCE   2  (bases 1 to 8678)
  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: 
            "10.1074/jbc.M807365200"; journalName: "J Biol Chem"; date: 
            "2009-04-10- 10"; volume: "284"; issue: "15"; pages: "9700-12"
COMMENT     Sequence Label: pEF-Bos-FLAG MDA5 MI
FEATURES             Location/Qualifiers
     source          1..8678
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..1179
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          232..1170
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1127..1147
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1196..1205
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1205..1228
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1232..1255
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(5250..5267)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5250..5266)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5259..5281)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5479..5934
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5566..5585)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5776..5797
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5951..5970)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6070..6092
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6130..6148)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6216..6320
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6321..6389,6390..7181)
                     /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     complement(6539..6558)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7352..7940
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7841..7860
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8094..8111
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8228..8249
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8264..8269,8270..8287,8288..8294)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8302..8318
                     /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     8307..8329
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8326..8342
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8326..8342
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        8374..8584
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        8396..8592
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      8443..8578
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
       61 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
      121 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
      181 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
      241 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
      301 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      361 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      421 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      481 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      541 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      601 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      661 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      721 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
      781 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
      841 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
      901 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
      961 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1021 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1081 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1141 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattctcta gataagccac
     1201 catggattat aaggatgatg atgataaagg tgattataag gatgatgatg ataaaggttc
     1261 gaatgggtat tccacagacg agaatttccg ctatctcatc tcgtgcttca gggccagggt
     1321 gaaaatgtac atccaggtgg agcctgtgct ggactacctg acctttctgc ctgcagaggt
     1381 gaaggagcag attcagagga cagtcgccac ctccgggaac atgcaggcag ttgaactgct
     1441 gctgagcacc ttggagaagg gagtctggca ccttggttgg actcgggaat tcgtggaggc
     1501 cctccggaga accggcagcc ctctggccgc ccgctacatg aaccctgagc tcacggactt
     1561 gccctctcca tcgtttgaga acgctcatga tgaatatctc caactgctga acctccttca
     1621 gcccactctg gtggacaagc ttctagttag agacgtcttg gataagtgca tggaggagga
     1681 actgttgaca attgaagaca gaaaccggat tgctgctgca gaaaacaatg gaaatgaatc
     1741 aggtgtaaga gagctactaa aaaggattgt gcagaaagaa aactggttct ctgcatttct
     1801 gaatgttctt cgtcaaacag gaaacaatga acttgtccaa gagttaacag gctctgattg
     1861 ctcagaaagc aatgcagaga ttgagaattt atcacaagtt gatggtcctc aagtggaaga
     1921 gcaacttctt tcaaccacag ttcagccaaa tctggagaag gaggtctggg gcatggagaa
     1981 taactcatca gaatcatctt ttgcagattc ttctgtagtt tcagaatcag acacaagttt
     2041 ggcagaagga agtgtcagct gcttagatga aagtcttgga cataacagca acatgggcag
     2101 tgattcaggc accatgggaa gtgattcaga tgaagagaat gtggcagcaa gagcatcccc
     2161 ggagccagaa ctccagctca ggccttacca aatggaagtt gcccagccag ccttggaagg
     2221 gaagaatatc atcatctgcc tccctacagg gagtggagca accagagtgg ctgtttacat
     2281 tgccaaggat cacttagaca agaagaaaaa agcatctgag cctggaaaag ttatagttct
     2341 tgtcaataag gtactgctag ttgaacagct cttccgcaag gagttccaac catttttgaa
     2401 gaaatggtat cgtgttattg gattaagtgg tgatacccaa ctgaaaatat catttccaga
     2461 agttgtcaag tcctgtgata ttattatcag tacagctcaa atccttgaaa actccctctt
     2521 aaacttggaa aatggagaag atgctggtgt tcaattgtca gacttttccc tcattatcat
     2581 tgatgaatgt catcacacca acaaagaagc agtgtataat aacatcatga ggcattattt
     2641 gatgcagaag ttgaaaaaca atagactcaa gaaagaaaac aaaccagtga ttccccttcc
     2701 tcagatactg ggactaacag cttcacctgg tgttggaggg gccacgaagc aagccaaagc
     2761 tgaagaacac attttaaaac tatgtgccaa tcttgatgca tttactatta aaactgttaa
     2821 agaaaacctt gatcaactga aaaaccaaat acaggagcca tgcaagaagt ttgccattgc
     2881 agatgcaacc agagaagatc catttaaaga gaaacttcta gaaataatga caaggattca
     2941 aacttattgt caaatgagtc caatgtcaga ttttggaact caaccctatg aacaatgggc
     3001 cattcaaatg gaaaaaaaag ctgcaaaaga aggaaatcgc aaagaacgtg tttgtgcaga
     3061 acatttgagg aagtacaatg aggccctaca aattaatgac acaattcgaa tgatagatgc
     3121 gtatactcat cttgaaactt tctataatga agagaaagat aagaagtttg cagtcataga
     3181 agatgatagt gatgagggtg gtgatgatga gtattgtgat ggtgataaag atgaggatga
     3241 tttaaagaaa cctttgaaac tggatgaaac agatagattt ctcatgactt tattttttga
     3301 aaacaataaa atgttgaaaa ggctggctga aaacccagaa tatgaaaatg aaaagctgac
     3361 caaattaaga aataccataa tggagcaata tactaggact gaggaatcag cacgaggaat
     3421 aatctttaca aaaacacgac agagtgcata tgcgctttcc cagtggatta ctgaaaatga
     3481 aaaatttgct gaagtaggag tcaaagccca ccatctgatt ggagctggac acagcagtga
     3541 gttcaaaccc atgacacaga atgaacaaaa agaagtcatt agtaaatttc gcactggaaa
     3601 aataaatctg cttatcgcta ccacagtggc agaagaaggt ctggatatta aagaatgtaa
     3661 cattgttatc cgttatggtc tcgtcaccaa tgaaatagcc atggtccagg cccgtggtcg
     3721 agccagagct gatgagagca cctacgtcct ggttgctcac agtggttcag gagttatcga
     3781 acatgagaca gttaatgatt tccgagagaa gatgatgtat aaagctatac attgtgttca
     3841 aaatatgaaa ccagaggagt atgctcataa gattttggaa ttacagatgc aaagtataat
     3901 ggaaaagaaa atgaaaacca agagaaatat tgccaagcat tacaagaata acccatcact
     3961 aataactttc ctttgcaaaa actgcagtgt gctagcctgt tctggggaag atatccatgt
     4021 aattgagaaa atgcatcacg tcaatatgac cccagaattc aaggaacttt acattgtaag
     4081 agaaaacaaa gcactgcaaa agaagtgtgc cgactatcaa ataaatggtg aaatcatctg
     4141 caaatgtggc caggcttggg gaacaatgat ggtgcacaaa ggcttagatt tgccttgtct
     4201 caaaataagg aattttgtag tggttttcaa aaataattca acaaagaaac aatacaaaaa
     4261 gtgggtagaa ttacctatca catttcccaa tcttgactat tcagaatgct gtttatttag
     4321 tgatgaggat tagcacttga ttgaatcgat gtcgacggta cctagagtga gggtccccac
     4381 ctgggaccct tgagagtatc aggtctccca cgtgggagac aagaaatccc tgtttaatat
     4441 ttaaacagca gtgttcccca tctgggtcct tgcacccctc actctggcct cagccgactg
     4501 cacagcggcc cctgcatccc cttggctgtg aggcccctgg acaagcagag gtggccagag
     4561 ctgggaggca tggccctggg gtcccacgaa tttgctgggg aatctcgttt ttcttcttaa
     4621 gacttttggg acatggtttg actcccgaac atcaccgacg tgtctcctgt ttttctgggt
     4681 ggcctcggga cacctgccct gcccccacga gggtcaggac tgtgactctt tttagggcca
     4741 ggcaggtgcc tggacatttg ccttgctgga cggggactgg ggatgtggga gggagcagac
     4801 aggaggaatc atgtcaggcc tgtgtgtgaa aggaagctcc actgtcaccc tccacctctt
     4861 caccccccac tcaccagtgt cccctccact gtcacattgt aactgaactt caggataata
     4921 aagtgtttgc ctccaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     4981 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     5041 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     5101 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     5161 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     5221 aaaaaaaaaa aaaaaaaaaa accgaattca ctggccgtcg ttttacaacg tcgtgactgg
     5281 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     5341 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     5401 gaatggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     5461 cgtcaaagca accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     5521 ttacgcgcag cgtgaccgct acacttgcca gcgccttagc gcccgctcct ttcgctttct
     5581 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     5641 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg
     5701 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     5761 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac tctatctcgg
     5821 gctattcttt tgatttataa gggattttgc cgatttcggt ctattggtta aaaaatgagc
     5881 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt
     5941 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa
     6001 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     6061 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     6121 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
     6181 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     6241 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     6301 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     6361 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     6421 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     6481 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     6541 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     6601 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     6661 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     6721 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     6781 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     6841 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     6901 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     6961 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     7021 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     7081 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     7141 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     7201 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     7261 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     7321 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     7381 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     7441 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     7501 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     7561 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     7621 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     7681 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     7741 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     7801 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     7861 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     7921 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     7981 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     8041 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     8101 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     8161 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     8221 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     8281 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     8341 accatgatta cgccaagctt aattccctcc ccagcaggca gaagtatgca aagcatgcat
     8401 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     8461 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     8521 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     8581 ggcttttgca aaaagctttg caaagatgga taaagtttta aacagagagg aatctttgca
     8641 gctaatggac cttctaggtc ttgaaaggag tgggaatt
//
