LOCUS       sequence-387841-        5999 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In vitro transcription/translation of HA-tagged wild-type Mettl3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5999)
  AUTHORS   Raj N, Wang M, Seoane JA, Zhao RL, Kaiser AM, Moonie NA, Demeter J, 
            Boutelle AM, Kerr CH, Mulligan AS, Moffatt C, Zeng SX, Lu H, Barna 
            M, Curtis C, Chang HY, Jackson PK, Attardi LD
  TITLE     The Mettl3 epitranscriptomic writer amplifies p53 stress responses.
  JOURNAL   Mol Cell. 2022 Jul 7;82(13):2370-2384.e10. doi: 
            10.1016/j.molcel.2022.04.010. Epub 2022 May 4.
  PUBMED    35512709
REFERENCE   2  (bases 1 to 5999)
  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.molcel.2022.04.010"; journalName: "Mol Cell"; date: 
            "2022-07-7- 7"; volume: "82"; issue: "13"; pages: "2370-2384.e10"
COMMENT     Sequence Label: pCS2-HA-Mettl3
FEATURES             Location/Qualifiers
     source          1..5999
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(528..1319,1320..1388))
                     /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     1151..1170
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1389..1493)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1519..1974)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1656..1677)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1868..1887
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2045..3029
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        3064..3082
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3064..3081
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             3120..3200
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     protein_bind    3225..3249
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(4996..5020)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    5080..5214
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5104..5123)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5158..5177
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5327..5347)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5327..5345)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5366..5382)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5366..5382)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5379..5401)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5390..5406
                     /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(5414..5420,5421..5438,5439..5444))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5459..5480
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5597..5614)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(5768..5999,1..357))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5848..5867)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
       61 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
      121 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
      181 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
      241 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
      301 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
      361 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
      421 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
      481 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
      541 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
      601 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
      661 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
      721 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
      781 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
      841 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
      901 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
      961 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     1021 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     1081 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     1141 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     1201 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     1261 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     1321 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     1381 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     1441 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     1501 ccccgaaaag tgccacctaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
     1561 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     1621 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     1681 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     1741 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     1801 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     1861 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     1921 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt
     1981 cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
     2041 gccagtcgac cgccaattca atatggcgta tatggactca tgccaattca atatggtgga
     2101 tctggacctg tgccaattca atatggcgta tatggactcg tgccaattca atatggtgga
     2161 tctggacccc agccaattca atatggcgga cttggcacca tgccaattca atatggcgga
     2221 cctggcactg tgccaactgg ggaggggtct acttggcacg gtgccaagtt tgaggagggg
     2281 tcttggccct gtgccaagtc cgccatattg aattggcatg gtgccaataa tggcggccat
     2341 attggctata tgccaggatc aatatatagg caatatccaa tatggcccta tgccaatatg
     2401 gctattggcc aggttcaata ctatgtattg gccctatgcc atatagtatt ccatatatgg
     2461 gttttcctat tgacgtagat agcccctccc aatgggcggt cccatatacc atatatgggg
     2521 cttcctaata ccgcccatag ccactccccc attgacgtca atggtctcta tatatggtct
     2581 ttcctattga cgtcatatgg gcggtcctat tgacgtatat ggcgcctccc ccattgacgt
     2641 caattacggt aaatggcccg cctggctcaa tgcccattga cgtcaatagg accacccacc
     2701 attgacgtca atgggatggc tcattgccca ttcatatccg ttctcacgcc ccctattgac
     2761 gtcaatgacg gtaaatggcc cacttggcag tacatcaata tctattaata gtaacttggc
     2821 aagtacatta ctattggaag tacgccaggg tacattggca gtactcccat tgacgtcaat
     2881 ggcggtaaat ggcccgcgat ggctgccaag tacatcccca ttgacgtcaa tggggagggg
     2941 caatgacgca aatgggcgtt ccattgacgt aaatgggcgg taggcgtgcc taatgggagg
     3001 tctatataag caatgctcgt ttagggaacc gccattctgc ctggggacgt cggagcaagc
     3061 ttgatttagg tgacactata gaatacaagc tacttgttct ttttgcagga tccaccatgt
     3121 acccatacga tgttccagat tacgcttatc cttatgacgt acctgactat gcataccctt
     3181 atgatgtacc agactacgct ggccggccag aattcaagga tcaaacaagt ttgtacaaaa
     3241 aagcaggctt aatgtcggac acgtggagct ctatccaggc ccataagaaa cagctggact
     3301 cgcttcgcga gagattgcag cggcgacgga agcaggactc tgggcacttg gatttaagga
     3361 acccagaagc ggcactgtcc ccaaccttcc gtagtgatag tcccgtgcct actgccccta
     3421 cctctagcgg ccctaagccc agcacaacat ctgtggcccc tgaactagct acagaccctg
     3481 agttagagaa gaagttgcta caccacctct cagatctggc cttgaccttg cccactgatg
     3541 ctgtttccat ccgtcttgcc atctctacgc cagatgcacc tgccactcaa gatggggtag
     3601 aaagccttct ccagaaattt gccgcccagg agttgattga ggtaaagcga ggtctcctcc
     3661 aagatgatgc acatcctact cttgtaactt acgctgacca ctccaagctg tctgccatga
     3721 tgggggctgt ggcagaaaag aaaggtcttg gagaggtagc agggaccatc gcagggcaga
     3781 aacggcgtgc agaacaggat ttgactacag tgaccacctt tgccagctct ttagcatctg
     3841 gtctggcctc ttcagcatca gaaccagcta aggagccggc taagaagtca aggaagcacg
     3901 ctgcctccga tgttgatctg gagatagaaa gccttttgaa ccaacagtca acgaaagaac
     3961 agcagagcaa gaaggtcagt caggagatcc tagagctatt aaataccaca acagccaagg
     4021 aacagtccat tgttgaaaag tttcgctctc gaggtcgggc ccaggtgcaa gaattttgtg
     4081 attatgggac caaggaagag tgcatgaaag ccagtgacgc tgaccggcct tgtcgcaagc
     4141 tgcacttcag acgaattatc aataagcaca ctgatgaatc tttaggtgac tgctctttcc
     4201 ttaacacatg tttccacatg gacacctgca aatatgttca ctatgaaatt gatgcttgtg
     4261 ttgattctga gagtcctggc agcaaggagc atatgccaag ccaggagctt gctcttacac
     4321 agagtgttgg gggtgactcc agtgctgatc gactctttcc acctcagtgg atctgttgtg
     4381 atatccgcta cctggacgtc agtatcttgg gcaaatttgc agttgtgatg gctgacccac
     4441 cttgggatat tcacatggag ctaccgtatg ggacattaac agatgatgag atgcgcaggc
     4501 tcaatatacc agtgctacag gatgacggct ttcttttcct ctgggtcaca ggaagggcca
     4561 tggaattggg cagagaatgt ctgaacctct ggggttatga acgggtggat gaaatcatct
     4621 gggtgaagac taatcagctg cagcgcatca ttaggacggg ccggacgggt cactggttaa
     4681 accacgggaa ggaacactgc ttggttggtg ttaagggaaa tcctcaagga ttcaaccagg
     4741 gcctggactg cgatgtgatt gtagctgagg ttcgttccac cagtcataaa ccagatgaaa
     4801 tatatggcat gattgagaga ctgtcccctg gcacccgcaa gattgagtta tttggacgac
     4861 cacacaatgt gcagcccaac tggattactc ttggaaacca actggatggg atacacctac
     4921 tagacccaga tgtggttgcc aggtttaagc aaaggtatcc ggacggcatc atctctaaac
     4981 ctaagaattt atagtaccca gctttcttgt acaaagtggt tcgatcctct cgagcctcta
     5041 gtggcgcgcc tctagaacta tagtgagtcg tattacgtag atccagacat gataagatac
     5101 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     5161 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
     5221 aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaattc
     5281 gcggccgcgg cgccaatgca ttgggcccgg tacccagctt ttgttccctt tagtgagggt
     5341 taattgcgcg cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     5401 tcacaattcc acacaacata cgagccggga gcataaagtg taaagcctgg ggtgcctaat
     5461 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     5521 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     5581 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     5641 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     5701 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     5761 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     5821 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     5881 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     5941 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtc
//
