LOCUS       sequence-111440-        6935 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for Bxb1-mediated recombination of the mCh-Dnmt3b6 cDNA into
            a MIN-tagged locus using Neomycin selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6935)
  AUTHORS   Mulholland CB, Smets M, Schmidtmann E, Leidescher S, Markaki Y, 
            Hofweber M, Qin W, Manzo M, Kremmer E, Thanisch K, Bauer C, Rombaut 
            P, Herzog F, Leonhardt H, Bultmann S
  TITLE     A modular open platform for systematic functional studies under 
            physiological conditions.
  JOURNAL   Nucleic Acids Res. 2015 May 24. pii: gkv550.
  PUBMED    26007658
REFERENCE   2  (bases 1 to 6935)
  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: "Nucleic 
            Acids Res. 2015 May 24. pii: gkv550."
COMMENT     Sequence Label: attB-mCh-Dnmt3b6-Poly(A)-NeoR
FEATURES             Location/Qualifiers
     source          1..6935
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        62..561
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(97..120)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     494..513
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             579..1373
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             579..1373
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1561..1580)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1605..1652
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     protein_bind    1823..1860
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     CDS             1863..2567
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(2005..2023)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2273..2292
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     regulatory      2613..2622
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2702..2713
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     polyA_signal    5050..5184
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5074..5093)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5128..5147
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        5320..5367
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5386..6189
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     rep_origin      complement(6318..6906)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6398..6417)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gatgccagct cattcctccc actcatgatc tatagatccc ccgggctgca ggaattctac
       61 cgggtagggg aggcgctttt cccaaggcag tctggagcat gcgctttagc agccccgctg
      121 ggcacttggc gctacacaag tggcctctgg cctcgcacac attccacatc caccggtagg
      181 cgccaaccgg ctccgttctt tggtggcccc ttcgcgccac cttctactcc tcccctagtc
      241 aggaagttcc cccccgcccc gcagctcgcg tcgtgcagga cgtgacaaat ggaagtagca
      301 cgtctcacta gtctcgtgca gatggacagc accgctgagc aatggaagcg ggtaggcctt
      361 tggggcagcg gccaatagca gctttgctcc ttcgctttct gggctcagag gctgggaagg
      421 ggtgggtccg ggggcgggct caggggcggg ctcaggggcg gggcgggcgc ccgaaggtcc
      481 tccggaggcc cggcattctg cacgcttcaa aagcgcacgt ctgccgcgct gttctcctct
      541 tcctcatctc cgggcctttc gaccgatcat caagcttaat gattgaacaa gatggattgc
      601 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
      661 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
      721 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat
      781 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
      841 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
      901 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
      961 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     1021 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     1081 ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc
     1141 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     1201 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     1261 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     1321 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     1381 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     1441 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     1501 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac
     1561 tgaaacacgg aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca
     1621 gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg
     1681 gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt
     1741 ttccccaccc caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc
     1801 ggcaagctta catcgagatc ccggcttgtc gacgacggcg gtctccgtcg tcaggatcat
     1861 ccgtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg cgcttcaagg
     1921 tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag ggcgagggcc
     1981 gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc cccctgccct
     2041 tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac gtgaagcacc
     2101 ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag tgggagcgcg
     2161 tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc ctgcaggacg
     2221 gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac ggccccgtaa
     2281 tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc gaggacggcg
     2341 ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac tacgacgctg
     2401 aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc tacaacgtca
     2461 acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa cagtacgaac
     2521 gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagagc ggcctgagga
     2581 gcagagccca ggcgagcaac agcgccgtgg acgccaccat gggcgatcgc atgaagggag
     2641 acagcagaca tctgaatgaa gaagagggtg ccagcgggta tgaggagtgc attatcgtta
     2701 atgggaactt cagtgaccag tcctcagaca cgaaggatgc tccctcaccc ccagtcttgg
     2761 aggcaatctg cacagagcca gtctgcacac cagagaccag aggccgcagg tcaagctccc
     2821 ggctgtctaa gagggaggtc tccagccttc tgaattacac gcaggacatg acaggagatg
     2881 gagacagaga tgatgaagta gatgatggga atggctctga tattctaatg ccaaagctca
     2941 cccgtgagac caaggacacc aggacgcgct ctgaaagccc ggctgtccga acccgacata
     3001 gcaatgggac ctccagcttg gagaggcaaa gagcctcccc cagaatcacc cgaggtcggc
     3061 agggccgcca ccatgtgcag gagtaccctg tggagtttcc ggctaccagg tctcggagac
     3121 gtcgagcatc gtcttcagca agcacgccat ggtcatcccc tgccagcgtc gacttcatgg
     3181 aagaagtgac acctaagagc gtcagtaccc catcagttga cttgagccag gatggagatc
     3241 aggagggtat ggataccaca caggtggatg cagagagcag agatggagac agcacagagt
     3301 atcaggatga taaagagttt ggaataggtg acctcgtgtg gggaaagatc aagggcttct
     3361 cctggtggcc tgccatggtg gtgtcctgga aagccacctc caagcgacag gccatgcccg
     3421 gaatgcgctg ggtacagtgg tttggtgatg gcaagttttc tgagatctct gctgacaaac
     3481 tggtggctct ggggctgttc agccagcact ttaatctggc taccttcaat aagctggttt
     3541 cttataggaa ggccatgtac cacactctgg agaaagccag ggttcgagct ggcaagacct
     3601 tctccagcag tcctggagag tcactggagg accagctgaa gcccatgctg gagtgggccc
     3661 acggtggctt caagcctact gggatcgagg gcctcaaacc caacaagaag caaccagtgg
     3721 ttaataagtc gaaggtgcgt cgttcagaca gtaggaactt agaacccagg agacgcgaga
     3781 acaaaagtcg aagacgcaca accaatgact ctgctgcttc tgagtccccc ccacccaagc
     3841 gcctcaagac aaatagctat ggcgggaagg accgagggga ggatgaggag agccgagaac
     3901 ggatggcttc tgaagtcacc aacaacaagg gcaatctgga agaccgctgt ttgtcctgtg
     3961 gaaagaagaa ccctgtgtcc ttccaccccc tctttgaggg tgggctctgt cagagttgcc
     4021 gggatcgctt cctagagctc ttctacatgt atgatgagga cggctatcag tcctactgca
     4081 ccgtgtgctg tgagggccgt gaactgctgc tgtgcagtaa cacaagctgc tgcagatgct
     4141 tctgtgtgga gtgtctggag gtgctggtgg gcgcaggcac agctgaggat gccaagctgc
     4201 aggaaccctg gagctgctat atgtgcctcc ctcagcgctg ccatggggtc ctccgacgca
     4261 ggaaagattg gaacatgcgc ctgcaagact tcttcactac tgatcctgac ctggaagaat
     4321 ttgagccacc caagttgtac ccagcaattc ctgcagccaa aaggaggccc attagagtcc
     4381 tgtctctgtt tgatggaatt gcaacggggt acttggtgct caaggagttg ggtattaaag
     4441 tggaaaagta cattgcctcc gaagtctgtg cagagtccat cgctgtggga actgttaagc
     4501 atgaaggcca gatcaaatat gtcaatgacg tccggaaaat caccaagaaa aatattgaag
     4561 agtggggccc gttcgacttg gtgattggtg gaagcccatg caatgatctc tctaacgtca
     4621 atcctgcccg caaaggttta tatgagggca caggaaggct cttcttcgag ttttaccact
     4681 tgctgaatta tacccgcccc aaggagggcg acaaccgtcc attcttctgg atgttcgaga
     4741 atgttgtggc catgaaagtg aatgacaaga aagacatctc aagattcctg gcatgtaacc
     4801 cagtgatgat cgatgccatc aaggtgtctg ctgctcacag ggcccggtac ttctggggta
     4861 acctacccgg aatgaacagg atcttcggct tccctgctca ctacacggac gtgtccaaca
     4921 tgggccgcgg cgcccgtcag aagctgctgg gcaggtcctg gagtgtaccg gtcatcagac
     4981 acctgtttgc ccccttgaag gactactttg cctgtgaata ggcggccgca gttaacgaat
     5041 tctctagagg atccagacat gataagatac attgatgagt ttggacaaac cacaactaga
     5101 atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc
     5161 attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt
     5221 cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtatggct
     5281 gattatgatc ctgcaagcct cgtcgccgcg gtttattctg ttgacaatta atcatcggca
     5341 tagtatatcg gcatagtata atacgacaag gtgaggaact aaaccatggg atcggccatt
     5401 gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat
     5461 gactgggcac aacagacgat cggctgctct gatgccgccg tgttccggct gtcagcgcag
     5521 gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac
     5581 gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac
     5641 gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc
     5701 ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg
     5761 ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag
     5821 cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat
     5881 caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag
     5941 gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc
     6001 ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg
     6061 ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg
     6121 ctttacggta tcgccgcccc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag
     6181 ttcttctgag cgggactctg gggttcgaat aaagaccgac caagcgacgt ctgagagctc
     6241 cctggcgaat tcggtaccaa taaaagagct ttattttcat gatctgtgtg ttggtttttg
     6301 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     6361 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     6421 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     6481 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     6541 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     6601 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     6661 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     6721 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     6781 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     6841 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     6901 tctcaagaag atcctttgat cttttctagt gtgcg
//
