LOCUS       sequence-171324-        6744 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses murine Mettl4 containing N-terminal 3XFlag tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6744)
  AUTHORS   Kweon SM, Chen Y, Moon E, Kvederaviciute K, Klimasauskas S, Feldman 
            DE
  TITLE     An Adversarial DNA N(6)-Methyladenine-Sensor Network Preserves 
            Polycomb Silencing.
  JOURNAL   Mol Cell. 2019 Apr 5. pii: S1097-2765(19)30222-9. doi: 
            10.1016/j.molcel.2019.03.018.
  PUBMED    30982744
REFERENCE   2  (bases 1 to 6744)
  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: "Mol Cell. 
            2019 Apr 5. pii: S1097-2765(19)30222-9. doi: 
            10.1016/j.molcel.2019.03.018."
COMMENT     Sequence Label: p3XFlag-Mettl4
FEATURES             Location/Qualifiers
     source          1..6744
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..66
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             join(1425..1427,1428..1430,1431..2147)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYKS"
     primer_bind     complement(1470..1491)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1731..1750)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2078..2099
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2155..2777
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2354..2373)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2801..2821)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2806..3135
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2986..3121
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3173..3192)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3174..3192)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3206..3222)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3206..3222)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3219..3241)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3230..3246
                     /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(3254..3260,3261..3278,3279..3284))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3299..3320
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3437..3454)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3608..4196)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3688..3707)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4367..5158,5159..5227))
                     /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     4990..5009
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5228..5332)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5359..5814
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5446..5465)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5656..5677
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5940..5962
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5954..5971
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5955..5971
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        6132..6511
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6512..6715
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6666..6686
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 gactacaaag accatgacgg tgattataaa gatcatgaca tcgattacaa ggatgacgat
       61 gacaagcttt ctgtggtgca tcatttgcct cctgggtggt tactggatca tctttctttc
      121 attaacaaag taaactatca actttgccag catcaagagt ctttttgtag taaaaacaac
      181 cctacttctt ctgtctacat ggattctctt cagctggatc ccgggtctcc ttttggagct
      241 cctgctatgt gctttgctcc tgacttcact actgtgtcgg gcaatgacga tgaaggaagt
      301 tgtgaagtga tcaccgaaaa atatgttttc cgatctgagc tatttaatgt caccaaacct
      361 tacatagttc cagctgttca caaagagcgc caacaaagta acaaaaacga aaatctagtg
      421 actgactata aacaagaggt ctctgtttct gttgggaaga tcagagagct gattttggat
      481 ggatcttcga agttaatcca agaaggtctc agaagtggtt ttctttaccc acttgttgaa
      541 aaacaggatg gaagtagcgg atgcatcact ttaccacttg atgcctgcaa tttatcagag
      601 ttatgtgaga tggcaaaaca tttgccttcc ttaaatgaaa tggagcttca aacgctacag
      661 ttgatgggag atgatgtgtc tgttatagag ctggatttat cctcacagat cattgaaaac
      721 aactccagct tttccaaaat gattacttta atgggacaaa aatacctact gccaccacaa
      781 agcagctttc ttttgtctga catttcttgt atgcagccac ttcttaactg tggtaagaca
      841 tttgatgcaa ttgtgattga tccaccatgg gagaacaaat cggttaaaag aagtaacagg
      901 tacagtagtt tatcaccaca gcagataaag cgaatgccta tccctaaact ggctgctgct
      961 gactgtctca ttgttacttg ggtgaccaac aggcagaaac atctgtgttt tgtgaaggag
     1021 gaactatatc cttcgtggtc tgtggaagtt gttgcagagt ggtactgggt aaaaatcaca
     1081 aattcaggag agtttgtgtt cccgttggat tctccacaca aaaagcctta tgaatgtctt
     1141 gtactgggga gagttaaaga aaaaactccc ttagcattaa ggaatccaga tgtaagaata
     1201 cccccagttc cagatcagaa gttgattgtc agtgtgccct gtgttcttca ctcacacaaa
     1261 ccgcctctta ccgaagttct gagagactac atcaagccag gtggccagtg cttggaattg
     1321 tttgctagga atttacagcc aggttggatg agttggggca acgaagtcct caagtttcag
     1381 cacatggatt atttcattgc tctggagtcc ggatgctgaa ttcaatggtg agcaagggcg
     1441 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     1501 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     1561 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccttcg
     1621 gctacggcct gcagtgcttc gcccgctacc ccgaccacat gaagcagcac gacttcttca
     1681 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     1741 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     1801 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     1861 acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact
     1921 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     1981 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agctaccagt
     2041 ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     2101 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtcctgagga tcccgggtgg
     2161 catccctgtg acccctcccc agtgcctctc ctggccctgg aagttgccac tccagtgccc
     2221 accagccttg tcctaataaa attaagttgc atcattttgt ctgactaggt gtccttctat
     2281 aatattatgg ggtggagggg ggtggtatgg agcaaggggc aagttgggaa gacaacctgt
     2341 agggcctgcg gggtctattg ggaaccaagc tggagtgcag tggcacaatc ttggctcact
     2401 gcaatctccg cctcctgggt tcaagcgatt ctcctgcctc agcctcccga gttgttggga
     2461 ttccaggcat gcatgaccag gctcagctaa tttttgtttt tttggtagag acggggtttc
     2521 accatattgg ccaggctggt ctccaactcc taatctcagg tgatctaccc accttggcct
     2581 cccaaattgc tgggattaca ggcgtgaacc actgctccct tccctgtcct tctgatttta
     2641 aaataactat accagcagga ggacgtccag acacagcata ggctacctgg ccatgcccaa
     2701 ccggtgggac atttgagttg cttgcttggc actgtcctct catgcgttgg gtccactcag
     2761 tagatgcctg ttgaattggg tacgcggcca gcttggctgt ggaatgtgtg tcagttaggg
     2821 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     2881 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     2941 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     3001 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     3061 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     3121 ctaggctttt gcaaaaagct cctcgaggaa ctgaaaaacc agaaagttaa ttccctatag
     3181 tgagtcgtat taaattcgta atcatgtcat agctgtttcc tgtgtgaaat tgttatccgc
     3241 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     3301 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     3361 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     3421 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     3481 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     3541 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     3601 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     3661 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     3721 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     3781 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     3841 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     3901 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     3961 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     4021 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     4081 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     4141 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     4201 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     4261 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     4321 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     4381 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     4441 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     4501 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     4561 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     4621 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     4681 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     4741 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     4801 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     4861 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     4921 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     4981 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     5041 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     5101 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     5161 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     5221 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     5281 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     5341 cccgaaaagt gccacctgac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     5401 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     5461 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     5521 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     5581 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     5641 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     5701 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     5761 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttccattc
     5821 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     5881 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
     5941 ccagtcacga cgttgtaaaa cgacggccag tgccaagctg atctatacat tgaatcaata
     6001 ttggcaatta gccatattag tcattggtta tatagcataa atcaatattg gctattggcc
     6061 attgcatacg ttgtatctat atcataatat gtacatttat attggctcat gtccaatatg
     6121 accgccatgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
     6181 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
     6241 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
     6301 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
     6361 ggcagtacat caagtgtatc atatgccaag tccgccccct attgacgtca atgacggtaa
     6421 atggcccgcc tggcattatg cccagtacat gaccttacgg gactttccta cttggcagta
     6481 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acaccaatgg
     6541 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
     6601 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaata accccgcccc
     6661 gttgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctcgttt
     6721 agtgaaccgt cagaattaac catg
//
