LOCUS       sequence-269997-        4970 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  KRAB-m6A Reader. Repress transcription near Gm6ATC sites. 
            pUBC-KRAB-NLS-DpnI(aa146-254).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4970)
  AUTHORS   Park M, Patel N, Keung AJ, Khalil AS
  TITLE     Engineering Epigenetic Regulation Using Synthetic Read-Write 
            Modules.
  JOURNAL   Cell. 2019 Jan 10;176(1-2):227-238.e20. doi: 
            10.1016/j.cell.2018.11.002. Epub 2018 Dec 6.
  PUBMED    30528434
REFERENCE   2  (bases 1 to 4970)
  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: "Cell."; 
            date: "2019-01-10"; volume: "176(1-2):227-238.e20. doi"; pages: " 
            10.1016/j.cell.2018.11.002. Epub 2018 Dec 6"
COMMENT     Sequence Label: pMP315
FEATURES             Location/Qualifiers
     source          1..4970
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(88..879,880..948))
                     /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     711..730
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(949..1053)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1097..2308
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     2145..2164
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     regulatory      2326..2335
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2362..2556
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQILYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEPWLV"
     CDS             2623..2643
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3092..3111)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3157..3212
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3211..3230)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3573..3589)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3573..3589)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3586..3608)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3597..3613
                     /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(3621..3627,3628..3645,3646..3651))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3666..3687
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      3772..3906
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3926..4060
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3963..3982)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4017..4036
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4128..4145)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4299..4887)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4379..4398)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
       61 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
      121 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
      181 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
      241 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
      301 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
      361 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
      421 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
      481 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
      541 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
      601 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
      661 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
      721 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
      781 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
      841 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
      901 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
      961 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     1021 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctgg
     1081 gtcgacattg attattgact agtgcgccgg gttttggcgc ctcccgcggg cgcccccctc
     1141 ctcacggcga gcgctgccac gtcagacgaa gggcgcagcg agcgtcctga tccttccgcc
     1201 cggacgctca ggacagcggc ccgctgctca taagactcgg ccttagaacc ccagtatcag
     1261 cagaaggaca ttttaggacg ggacttgggt gactctaggg cactggtttt ctttccagag
     1321 agcggaacag gcgaggaaaa gtagtccctt ctcggcgatt ctgcggaggg atctccgtgg
     1381 ggcggtgaac gccgatgatt atataaggac gcgccgggtg tggcacagct agttccgtcg
     1441 cagccgggat ttgggtcgcg gttcttgttt gtggatcgct gtgatcgtca cttggtgagt
     1501 agcgggctgc tgggctggcc ggggctttcg tggccgccgg gccgctcggt gggacggaag
     1561 cgtgtggaga gaccgccaag ggctgtagtc tgggtccgcg agcaaggttg ccctgaactg
     1621 ggggttgggg ggagcgcagc aaaatggcgg ctgttcccga gtcttgaatg gaagacgctt
     1681 gtgaggcggg ctgtgaggtc gttgaaacaa ggtggggggc atggtgggcg gcaagaaccc
     1741 aaggtcttga ggccttcgct aatgcgggaa agctcttatt cgggtgagat gggctggggc
     1801 accatctggg gaccctgacg tgaagtttgt cactgactgg agaactcggt ttgtcgtctg
     1861 ttgcgggggc ggcagttatg gcggtgccgt tgggcagtgc acccgtacct ttgggagcgc
     1921 gcgccctcgt cgtgtcgtga cgtcacccgt tctgttggct tataatgcag ggtggggcca
     1981 cctgccggta ggtgtgcggt aggcttttct ccgtcgcagg acgcagggtt cgggcctagg
     2041 gtaggctctc ctgaatcgac aggcgccgga cctctggtga ggggagggat aagtgaggcg
     2101 tcagtttctt tggtcggttt tatgtaccta tcttcttaag tagctgaagc tccggttttg
     2161 aactatgcgc tcggggttgg cgagtgtgtt ttgtgaagtt ttttaggcac cttttgaaat
     2221 gtaatcattt gggtcaatat gtaattttca gtgttagact agtaaattgt ccgctaaatt
     2281 ctggccgttt ttggcttttt tgttagacac gcgtttaatt aagccgccac catggatgct
     2341 aagtcactga ctgcctggtc ccggacactg gtgaccttca aggatgtgtt tgtggacttc
     2401 accagggagg agtggaagct gctggacact gctcagcaga tcctgtacag aaatgtgatg
     2461 ctggagaact ataagaacct ggtttccttg ggttatcagc ttactaagcc agatgtgatc
     2521 ctccggttgg agaagggaga agagccctgg ctggtggaga gagaaattca ccaagagacc
     2581 catcctgatt cagagactgc atttgaaatc aaatcatcag ttccgaaaaa gaaacgcaaa
     2641 gtcctcgagg gaggcggtgg aagcggcacc tgcaggtccc aggtgccatc taaaggccgg
     2701 atctttctcg tccaagatgg ccaggttagg gatccagaga aagtgacaaa agagttcaaa
     2761 cagggactct ttctccggaa gtcatccctg tcctccaggg ggtggacaat cgagatcctg
     2821 aactgcattg acaaaatcga gggctccgag tttacgcttg aggacatgta ccgattcgag
     2881 tctgacctta agaatatttt cgtgaagaac aatcacatca aagaaaaaat ccgccagcag
     2941 ctgcagatcc tcagagacaa agaaatcatc gagttcaaag gtaggggtaa gtaccgcaag
     3001 ctgtaagcgg ccgcactcct caggtgcagg ctgcctatca gaaggtggtg gctggtgtgg
     3061 ccaatgccct ggctcacaaa taccactgag atctttttcc ctctgccaaa aattatgggg
     3121 acatcatgaa gccccttgag catctgactt ctggctaata aaggaaattt attttcattg
     3181 caatagtgtg ttggaatttt ttgtgtctct cactcggaag gacatatggg agggcaaatc
     3241 atttaaaaca tcagaatgag tatttggttt agagtttggc aacatatgcc catatgctgg
     3301 ctgccatgaa caaaggttgg ctataaagag gtcatcagta tatgaaacag ccccctgctg
     3361 tccattcctt attccataga aaagccttga cttgaggtta gatttttttt atattttgtt
     3421 ttgtgttatt tttttcttta acatccctaa aattttcctt acatgtttta ctagccagat
     3481 ttttcctcct ctcctgacta ctcccagtca tagctgtccc tcttctctta tggagatccc
     3541 tcgacctgca gcccaagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     3601 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     3661 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     3721 ggaaacctgt cgtgccagcg gatccgcatc tcaattagtc agcaaccata gtcccgcccc
     3781 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     3841 cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg
     3901 gaggcctagg cttttgcaaa aagctaactt gtttattgca gcttataatg gttacaaata
     3961 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     4021 tttgtccaaa ctcatcaatg tatcttatca tgtctggatc cgctgcatta atgaatcggc
     4081 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     4141 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     4201 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     4261 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     4321 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     4381 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     4441 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     4501 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4561 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     4621 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     4681 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     4741 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     4801 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     4861 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     4921 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
//
