LOCUS       sequence-401795-        9340 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral expression of Flag-Tagged human METTL3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9340)
  TITLE     Alarcon lab plasmids
REFERENCE   2  (bases 1 to 9340)
  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"
COMMENT     Sequence Label:  pLenti-C-Myc-DDK-IRES-Puro METTL3
FEATURES             Location/Qualifiers
     source          1..9340
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(77..736)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     651..670
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(737..841)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(916..935)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1106..1485
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1487..1685
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             1703..1883
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1930..2055
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    2552..2785
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2970..3014
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    3312..3429
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        3512..3891
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3892..4095
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4121..4140
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4121..4139
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5995..6024
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             6043..6066
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    6100..6673
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    6123..6675
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(6267..6284)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     6494..6513
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             6675..7274
                     /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(6675..6694)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7171..7191
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    complement(7304..7337)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    7393..7981
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7446..7466)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7864..7875)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(7984..8000)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7985..8001)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             8510..8690
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(8752..9340)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8832..8851)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
       61 gggattttgg tcatgattac gccccgccct gccactcatc gcagtactgt tgtaattcat
      121 taagcattct gccgacatgg aagccatcac aaacggcatg atgaacctga atcgccagcg
      181 gcatcagcac cttgtcgcct tgcgtataat atttgcccat ggtgaaaacg ggggcgaaga
      241 agttgtccat attggccacg tttaaatcaa aactggtgaa actcacccag ggattggctg
      301 agacgaaaaa catattctca ataaaccctt tagggaaata ggccaggttt tcaccgtaac
      361 acgccacatc ttgcgaatat atgtgtagaa actgccggaa atcgtcgtgg tattcactcc
      421 agagcgatga aaacgtttca gtttgctcat ggaaaacggt gtaacaaggg tgaacactat
      481 cccatatcac cagctcaccg tctttcattg ccatacggaa ctccggatga gcattcatca
      541 ggcgggcaag aatgtgaata aaggccggat aaaacttgtg cttatttttc tttacggtct
      601 ttaaaaaggc cgtaatatcc agctgaacgg tctggttata ggtacattga gcaactgact
      661 gaaatgcctc aaaatgttct ttacgatgcc attgggatat atcaacggtg gtatatccag
      721 tgattttttt ctccatactc ttcctttttc aatattattg aagcatttat cagggttatt
      781 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
      841 gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc ccgatcccct
      901 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atctgctccc
      961 tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca
     1021 aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg cgctgcttcg
     1081 cgatgtacgg gccagatatc gcgttgacat tgattattga ctagttatta atagtaatca
     1141 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
     1201 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
     1261 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
     1321 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
     1381 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
     1441 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg
     1501 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
     1561 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
     1621 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
     1681 agcagcgcgt tttgcctgta ctgggtctct ctggttagac cagatctgag cctgggagct
     1741 ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca
     1801 agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta
     1861 gtcagtgtgg aaaatctcta gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa
     1921 ccagaggagc tctctcgacg caggactcgg cttgctgaag cgcgcacggc aagaggcgag
     1981 gggcggcgac tggtgagtac gccaaaaatt ttgactagcg gaggctagaa ggagagagat
     2041 gggtgcgaga gcgtcagtat taagcggggg agaattagat cgcgatggga aaaaattcgg
     2101 ttaaggccag ggggaaagaa aaaatataaa ttaaaacata tagtatgggc aagcagggag
     2161 ctagaacgat tcgcagttaa tcctggcctg ttagaaacat cagaaggctg tagacaaata
     2221 ctgggacagc tacaaccatc ccttcagaca ggatcagaag aacttagatc attatataat
     2281 acagtagcaa ccctctattg tgtgcatcaa aggatagaga taaaagacac caaggaagct
     2341 ttagacaaga tagaggaaga gcaaaacaaa agtaagacca ccgcacagca agcggccggc
     2401 cgctgatctt cagacctgga ggaggagata tgagggacaa ttggagaagt gaattatata
     2461 aatataaagt agtaaaaatt gaaccattag gagtagcacc caccaaggca aagagaagag
     2521 tggtgcagag agaaaaaaga gcagtgggaa taggagcttt gttccttggg ttcttgggag
     2581 cagcaggaag cactatgggc gcagcgtcaa tgacgctgac ggtacaggcc agacaattat
     2641 tgtctggtat agtgcagcag cagaacaatt tgctgagggc tattgaggcg caacagcatc
     2701 tgttgcaact cacagtctgg ggcatcaagc agctccaggc aagaatcctg gctgtggaaa
     2761 gatacctaaa ggatcaacag ctcctgggga tttggggttg ctctggaaaa ctcatttgca
     2821 ccactgctgt gccttggaat gctagttgga gtaataaatc tctggaacag atttggaatc
     2881 acacgacctg gatggagtgg gacagagaaa ttaacaatta cacaagctta atacactcct
     2941 taattgaaga atcgcaaaac cagcaagaaa agaatgaaca agaattattg gaattagata
     3001 aatgggcaag tttgtggaat tggtttaaca taacaaattg gctgtggtat ataaaattat
     3061 tcataatgat agtaggaggc ttggtaggtt taagaatagt ttttgctgta ctttctatag
     3121 tgaatagagt taggcaggga tattcaccat tatcgtttca gacccacctc ccaaccccga
     3181 ggggacccga caggcccgaa ggaatagaag aagaaggtgg agagagagac agagacagat
     3241 ccattcgatt agtgaacgga tcggcactgc gtgcgccaat tctgcagaca aatggcagta
     3301 ttcatccaca attttaaaag aaaagggggg attggggggt acagtgcagg ggaaagaata
     3361 gtagacataa tagcaacaga catacaaact aaagaattac aaaaacaaat tacaaaaatt
     3421 caaaattttc gggtttatta cagggacagc agagatccag tttggttagt accgggcccg
     3481 ctctagacat gtccaatatg accgccatgt tgacattgat tattgactag ttattaatag
     3541 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
     3601 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
     3661 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
     3721 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tccgccccct
     3781 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttacgg
     3841 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
     3901 ttttggcagt acaccaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
     3961 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
     4021 tgtcgtaata accccgcccc gttgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
     4081 tatataagca gagctcgttt agtgaaccgt cagaattttg taatacgact cactataggg
     4141 cggccgggaa ttcgtcgact ggatccggta ccgaggagat ctgccgccgc gatcgccggc
     4201 gcgccagatc tcaagcttaa ctagctagcg gaccgacgcg tacgcggccg catgtcggac
     4261 acgtggagct ctatccaggc ccacaagaag cagctggact ctctgcggga gaggctgcag
     4321 cggaggcgga agcaggactc ggggcacttg gatctacgga atccagaggc agcattgtct
     4381 ccaaccttcc gtagtgacag cccagtgcct actgcaccca cctctggtgg ccctaagccc
     4441 agcacagctt cagcagttcc tgaattagct acagatcctg agttagagaa gaagttgcta
     4501 caccacctct ctgatctggc cttaacattg cccactgatg ctgtgtccat ctgtcttgcc
     4561 atctccacgc cagatgctcc tgccactcaa gatggggtag aaagcctcct gcagaagttt
     4621 gcagctcagg agttgattga ggtaaagcga ggtctcctac aagatgatgc acatcctact
     4681 cttgtaacct atgctgacca ttccaagctc tctgccatga tgggtgctgt ggcagaaaag
     4741 aagggccctg gggaggtagc agggactgtc acagggcaga agcggcgtgc agaacaggac
     4801 tcgactacag tagctgcctt tgccagttcg ttagtctctg gtctgaactc ttcagcatcg
     4861 gaaccagcaa aggagccagc caagaaatca aggaaacatg ctgcctcaga tgttgatctg
     4921 gagatagaga gccttctgaa ccaacagtcc actaaggaac aacagagcaa gaaggtcagt
     4981 caggagatcc tagagctatt aaatactaca acagccaagg aacaatccat tgttgaaaaa
     5041 tttcgctctc gaggtcgggc ccaagtgcaa gaattctgtg actatggaac caaggaggag
     5101 tgcatgaaag ccagtgatgc tgatcgaccc tgtcgcaagc tgcacttcag acgaattatc
     5161 aataaacaca ctgatgagtc tttaggtgac tgctctttcc ttaatacatg tttccacatg
     5221 gatacctgca agtatgttca ctatgaaatt gatgcttgca tggattctga ggcccctggc
     5281 agcaaagacc acacgccaag ccaggagctt gctcttacac agagtgtcgg aggtgattcc
     5341 agtgcagacc gactcttccc acctcagtgg atctgttgtg atatccgcta cctggacgtc
     5401 agtatcttgg gcaagtttgc agttgtgatg gctgacccac cctgggatat tcacatggaa
     5461 ctgccctatg ggaccctgac agatgatgag atgcgcaggc tcaacatacc cgtactacag
     5521 gatgatggct ttctcttcct ctgggtcaca ggcagggcca tggagttggg gagagaatgt
     5581 ctaaatctct gggggtatga acgggtagat gaaattattt gggtgaagac aaatcaactg
     5641 caacgcatca ttcggacagg ccgtacaggt cactggttga accatgggaa ggaacactgc
     5701 ttggttggtg tcaaaggaaa tccccaaggc ttcaaccagg gtctggattg tgatgtgatc
     5761 gtagctgagg ttcgttccac cagtcataaa ccagatgaaa tctatggcat gattgaaaga
     5821 ctatctcctg gcactcgcaa gattgagtta tttggacgac cacacaatgt gcaacccaac
     5881 tggatcaccc ttggaaacca actggatggg atccacctac tagacccaga tgtggttgca
     5941 cggttcaagc aaaggtaccc agatggtatc atctctaaac ctaagaattt actcgagcag
     6001 aaactcatct cagaagagga tctggcagca aatgatatcc tggattacaa ggatgacgac
     6061 gataaggttt aaacctaggc gtagcggccg caaattccgc ccctctccct cccccccccc
     6121 taacgttact ggccgaagcc gcttggaata aggccggtgt gcgtttgtct atatgttatt
     6181 ttccaccata ttgccgtctt ttggcaatgt gagggcccgg aaacctggcc ctgtcttctt
     6241 gacgagcatt cctaggggtc tttcccctct cgccaaagga atgcaaggtc tgttgaatgt
     6301 cgtgaaggaa gcagttcctc tggaagcttc ttgaagacaa acaacgtctg tagcgaccct
     6361 ttgcaggcag cggaaccccc cacctggcga caggtgcctc tgcggccaaa agccacgtgt
     6421 ataagataca cctgcaaagg cggcacaacc ccagtgccac gttgtgagtt ggatagttgt
     6481 ggaaagagtc aaatggctct cctcaagcgt attcaacaag gggctgaagg atgcccagaa
     6541 ggtaccccat tgtatgggat ctgatctggg gcctcggtgc acatgcttta catgtgttta
     6601 gtcgaggtta aaaaaacgtc taggcccccc gaaccacggg gacgtggttt tcctttgaaa
     6661 aacacgatga taatatgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg
     6721 tcccccgggc agtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca
     6781 ccgtcgatcc agaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc
     6841 gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct
     6901 ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg
     6961 ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc
     7021 accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg
     7081 gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg
     7141 tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag cggctcggct
     7201 tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca
     7261 agcccggtgc ctgatgtaca agtaggattc gtcgagggac ctaataactt cgtatagcat
     7321 acattatacg aagttataca tgtttaaggg ttccggttcc actaggtaca attcgatatc
     7381 aagcttatcg ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt
     7441 aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct
     7501 attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt
     7561 tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac
     7621 gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct
     7681 ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca
     7741 ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt
     7801 ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc
     7861 ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct
     7921 cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg
     7981 catcgatacc gtcgacctcg atcgagacct agaaaaacat ggagcaatca caagtagcaa
     8041 tacagcagct accaatgctg attgtgcctg gctagaagca caagaggagg aggaggtggg
     8101 ttttccagtc acacctcagg tacctttaag accaatgact tacaaggcag ctgtagatct
     8161 tagccacttt ttaaaagaaa aggggggact ggaagggcta attcactccc aacgaagaca
     8221 agatatcctt gatctgtgga tctaccacac acaaggctac ttccctgatt ggcagaacta
     8281 cacaccaggg ccagggatca gatatccact gacctttgga tggtgctaca agctagtacc
     8341 agttgagcaa gagaaggtag aagaagccaa tgaaggagag aacacccgct tgttacaccc
     8401 tgtgagcctg catgggatgg atgacccgga gagagaagta ttagagtgga ggtttgacag
     8461 ccgcctagca tttcatcaca tggcccgaga gctgcatccg gactgtactg ggtctctctg
     8521 gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc
     8581 tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg
     8641 taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca gcatgtgagc
     8701 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     8761 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     8821 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     8881 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     8941 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     9001 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     9061 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     9121 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     9181 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     9241 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     9301 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
//
