LOCUS       sequence-210225-        6240 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty TALE vector driven by EF1a promoter, no Puro, contains DNMT3a 
            catalytic domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6240)
  AUTHORS   Pflueger C, Tan D, Swain T, Nguyen TV, Pflueger J, Nefzger C, Polo 
            JM, Ford E, Lister R
  TITLE     A modular dCas9-SunTag DNMT3A epigenome editing system overcomes 
            pervasive off-target activity of direct fusion dCas9-DNMT3A 
            constructs.
  JOURNAL   Genome Res. 2018 Jun 15. pii: gr.233049.117. doi: 
            10.1101/gr.233049.117.
  PUBMED    29907613
REFERENCE   2  (bases 1 to 6240)
  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: "Genome Res.
            2018 Jun 15. pii: gr.233049.117. doi: 10.1101/gr.233049.117."
COMMENT     Sequence Label: LLP245 pEF1a-TALE-DNMT3a-Wt
FEATURES             Location/Qualifiers
     source          1..6240
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        281..1459
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          512..1450
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1407..1427
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             1475..1501
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1505..1531
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1535..1561
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1571..1591
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2303..2323
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2366..3298
                     /codon_start=1
                     /product="catalytic domain of DNA methyltransferase 3A"
                     /label=DNMT3A catalytic domain
                     /translation="PSRLQMFFANNHDQEFDPPKVYPPVPAEKRKPIRVLSLFDGIATG
                     LLVLKDLGIQVDRYIASEVCEDSITVGMVRHQGKIMYVGDVRSVTQKHIQEWGPFDLVI
                     GGSPCNDLSIVNPARKGLYEGTGRLFFEFYRLLHDARPKEGDDRPFFWLFENVVAMGVS
                     DKRDISRFLESNPVMIDAKEVSAAHRARYFWGNLPGMNRPLASTVNDKLELQECLEHGR
                     IAKFSKVRTITTRSNSIKQGKDQHFPVFMNEKEDILWCTEMERVFGFPVHYTDVSNMSR
                     LARQRLLGRSWSVPVIRHLFAPLKEYFACV"
     intron          3452..3681
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     3664..3683
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     polyA_signal    3860..3981
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3897..3916)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4083..4099)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4083..4099)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4096..4118)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4107..4123
                     /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(4131..4137,4138..4155,4156..4161))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4176..4197
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4314..4331)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4485..5073)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4565..4584)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5244..6035,6036..6104))
                     /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     5867..5886
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6105..6209)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg tgcaaagatg gataaagttt taaacagaga ggaatctttg
      241 cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg cccgtcagtg
      301 ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgaac
      361 cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg
      421 cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct
      481 ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt cccgcgggcc
      541 tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct ggctgcagta
      601 cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga ggccttgcgc
      661 ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct ggggccgccg
      721 cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt ctctagccat
      781 ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc ttgtaaatgc
      841 gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg acggggcccg
      901 tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac cgagaatcgg
      961 acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc cgccgtgtat
     1021 cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag cggaaagatg
     1081 gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct cgggagagcg
     1141 ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg tcgcttcatg
     1201 tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga gcttttggag
     1261 tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc acactgagtg
     1321 ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg aatttgccct
     1381 ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa gtttttttct
     1441 tccatttcag gtgtcgtgac gtacggccac catgtacccc tatgacgtgc ctgattacgc
     1501 cgcttatcct tacgatgtgc ccgactatgc agcctaccca tacgatgtgc ccgactacgc
     1561 cgcaatggcc ccaaagaaga agcggaaggt cggtatccac ggagtcccag cagccgtaga
     1621 tttgagaact ttgggatatt cacagcagca gcaggaaaag atcaagccca aagtgaggtc
     1681 gacagtcgcg cagcatcacg aagccctggt gggtcatggg tttacacatg cccacatcgt
     1741 agccttgtcg cagcaccctg cagcccttgg cacggtcgcc gtcaagtacc aggacatgat
     1801 tgcggcgttg ccggaagcca cacatgaggc gatcgtcggt gtggggaaac agtggagcgg
     1861 agcccgcgcc cttgaggccc tgttgacggt cgcgggagag ctgagagggc ctccccttca
     1921 gctggacacg ggccagttgc tgaagatcgc gaagcgggga ggagtcacgg cggtcgaggc
     1981 ggtgcacgcg tggcgcaatg cgctcacggg agcacccctc aacctgacag agacgctggc
     2041 tgatgggcgt ctcggaggca gacccgcact ggagtcaatt gtggcccagc tttcgaggcc
     2101 ggaccccgcg ctggccgcac tcactaatga tcatcttgta gcgctggcct gcctcggcgg
     2161 acgacccgcc ttggatgcgg tgaagaaggg gctcccgcac gcgcctgcat tgattaagcg
     2221 gaccaacaga aggattcccg agaggacatc acatcgagtg gcaggtggcg gagggtctgg
     2281 gggaggcggt agtcaggcca gcccgaaaaa gaaaagaaaa gtgggacggg cgaaaggagg
     2341 cggtggctca ggcggtgggg gaagcccctc ccggctccag atgttcttcg ctaataacca
     2401 cgaccaggaa tttgaccctc caaaggttta cccacctgtc ccagctgaga agaggaagcc
     2461 catccgggtg ctgtctctct ttgatggaat cgctacaggg ctcctggtgc tgaaggactt
     2521 gggcattcag gtggaccgct acattgcctc ggaggtgtgt gaggactcca tcacggtggg
     2581 catggtgcgg caccagggga agatcatgta cgtcggggac gtccgcagcg tcacacagaa
     2641 gcatatccag gagtggggcc cattcgatct ggtgattggg ggcagtccct gcaatgacct
     2701 ctccatcgtc aaccctgctc gcaagggcct ctacgagggc actggccggc tgttctttga
     2761 gttctaccgc ctcctgcatg atgcgcggcc caaggaggga gatgatcgcc ccttcttctg
     2821 gctctttgag aatgtggtgg ccatgggcgt tagtgacaag agggacatct cgcgatttct
     2881 cgagtccaac cctgtgatga ttgatgccaa agaagtgtca gctgcacaca gggcccgcta
     2941 cttctggggt aaccttcccg gtatgaacag gccgttggca tccactgtga atgataagct
     3001 ggagctgcag gagtgtctgg agcatggcag gatagccaag ttcagcaaag tgaggaccat
     3061 tactacgagg tcaaactcca taaagcaggg caaagaccag cattttcctg tcttcatgaa
     3121 tgagaaagag gacatcttat ggtgcactga aatggaaagg gtatttggtt tcccagtcca
     3181 ctatactgac gtgtccaaca tgagccgctt ggcgaggcag agactgctgg gccggtcatg
     3241 gagcgtgcca gtcatccgcc acctcttcgc tccgctgaag gagtattttg cgtgtgtgga
     3301 ggtgcacacc aaccaagatc cgcttgatgc tgaagttcat acaaaccagg atcccctcga
     3361 tgccgaagtc catactaatc aggacccact ggacgcatga gggggaggct aactggaatt
     3421 aattcgctgt ctgcgagggc cagctgttgg ggtgagtact ccctctcaaa agcgggcatg
     3481 acttctgcgc taagattgtc agtttccaaa aacgaggagg atttgatatt cacctggccc
     3541 gcggtgatgc ctttgagggt ggccgcgtcc atctggtcag aaaagacaat ctttttgttg
     3601 tcaagcttga ggtgtggcag gcttgagatc tggccataca cttgagtgac aatgacatcc
     3661 actttgcctt tctctccaca ggtgtccact cccaggtcca actgcaggtc gatttaaatg
     3721 ggaattccac agcggccgca tgtctagata actgatcata atcagccata ccacatttgt
     3781 agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat
     3841 gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa
     3901 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     3961 caaactcatc aatgtatctt aatgcgtcga gtgcattcta gttgtggttt gtccaaactc
     4021 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca
     4081 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     4141 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     4201 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     4261 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     4321 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     4381 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     4441 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4501 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4561 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4621 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4681 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4741 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4801 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4861 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4921 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4981 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     5041 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     5101 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     5161 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     5221 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     5281 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     5341 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     5401 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5461 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5521 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5581 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5641 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5701 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5761 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5821 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5881 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5941 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     6001 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     6061 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     6121 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     6181 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
//
