LOCUS       sequence-232515-        4326 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient transfection; Expresses FRB-RBM38; CAGGS promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4326)
  AUTHORS   Du M, Jillette N, Zhu JJ, Li S, Cheng AW
  TITLE     CRISPR artificial splicing factors.
  JOURNAL   Nat Commun. 2020 Jun 12;11(1):2973. doi: 10.1038/s41467-020-16806-4.
  PUBMED    32532987
REFERENCE   2  (bases 1 to 4326)
  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: "Nat 
            Commun."; date: "2020-06-12"; volume: "11(1):2973. doi"; pages: " 
            10.1038/s41467-020-16806-4"
COMMENT     Sequence Label: pAC1810_pmax-FRB-RBM38
FEATURES             Location/Qualifiers
     source          1..4326
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        140..519
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        520..723
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     674..694
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          815..947
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     protein_bind    962..986
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      1004..1013
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1013..1078
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1103..1123
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1127..1147
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1151..1171
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1253..1522
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="EMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTLKETS
                     FNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISKQ"
     protein_bind    complement(2357..2381)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2434..2475
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(2501..2520)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    2530..2611
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     2555..2574
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(2809..3397)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2889..2908)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3478..4272)
                     /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             complement(3478..4272)
                     /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(3589..3608)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4199..4218
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 tgtcaatatt ggccattagc catattattc attggttata tagcataaat caatattggc
       61 tattggccat tgcatacgtt gtatctatat cataatatgt acatttatat tggctcatgt
      121 ccaatatgac cgccatgttg gcattgatta ttgactagtt attaatagta atcaattacg
      181 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
      241 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
      301 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
      361 gcccacttgg cagtacatca agtgtatcat atgccaagtc cgccccctat tgacgtcaat
      421 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttacggga ctttcctact
      481 tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac
      541 accaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
      601 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaataac
      661 cccgccccgt tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga
      721 ggtcgtttag tgaaccgtca gatcactagt agctttattg cggtagttta tcacagttaa
      781 attgctaacg cagtcagtgc tcgactgatc acaggtaagt atcaaggtta caagacaggt
      841 ttaaggaggc caatagaaac tgggcttgtc gagacagaga agattcttgc gtttctgata
      901 ggcacctatt ggtcttactg acatccactt tgcctttctc tccacaggga aaaaacaatt
      961 gacaagtttg tacaaaaaag caggctccga attcaccggt gccgccacca tggactacaa
     1021 ggatcacgac ggtgactata aggatcatga catcgactat aaggacgatg acgataagat
     1081 cgatggcgga ggcggatctg atccaaaaaa gaagagaaag gtagatccaa aaaagaagag
     1141 aaaggtagat ccaaaaaaga agagaaaggt aggatctacc ggatctagaa acgatggtgg
     1201 tggtggaagc gggggtgggg gcagcggtgg agggggaagc gggcgcgcca tggagatgtg
     1261 gcatgaaggc ctggaagagg catctcgttt gtactttggg gaaaggaacg tgaaaggcat
     1321 gtttgaggtg ctggagccct tgcatgctat gatggaacgg ggcccccaga ctctgaagga
     1381 aacatccttt aatcaggcct atggtcgaga tttaatggag gcccaagagt ggtgcaggaa
     1441 gtacatgaaa tcagggaatg tcaaggacct cacccaagcc tgggacctct attatcatgt
     1501 gttccgacga atctcaaagc agcagatctc gtacgcgtcc cggggcggtg gctcatctgg
     1561 cggaggtgga ggcggcggta gcggaggagg cgggtccggc ggcggcggta gtgggccggc
     1621 catgcttctg caaccggcgc catgtgcacc ttcagcaggt tttccaagac cgcttgctgc
     1681 gccaggcgct atgcacggca gccaaaaaga caccacgttt acaaagatat tcgtcggtgg
     1741 actcccttat catactactg acgcgtccct tagaaagtac ttcgagggtt tcggggatat
     1801 tgaagaagcc gtcgtcataa cggatcggca aactggaaag agccggggct acggttttgt
     1861 cacgatggct gatagggcag cggcagagag ggcttgtaaa gacccaaacc ctattatcga
     1921 tgggcgcaag gcaaacgtaa acctggcata ccttggggca aagccgcgct cattgcagac
     1981 tggcttcgct atcggcgtac aacaactcca cccaacattg attcaaagaa cttatggact
     2041 cacaccacat tacatttatc ccccagcaat agtacaacca tcagtcgtca ttcctgcagc
     2101 tcccgttcca tcactctcca gtccttatat agagtataca ccagcctctc cagcttatgc
     2161 ccaatacccc cctgccacct atgaccaata cccatacgcc gcgagcccgg ccaccgcagc
     2221 cagctttgta ggttatagtt acccggctgc cgttccccaa gctctcagcg ctgccgctcc
     2281 tgctggtact accttcgtac aataccaggc tcctcagttg cagcccgaca gaatgcaatg
     2341 attaattaag aattcgaccc agctttcttg tacaaagtgg ttgatatcca gcacagtggc
     2401 ggccgctcga gtctagaggg cccgcggttc gaaggtaagc ctatccctaa ccctctcctc
     2461 ggtctcgatt ctacgcgtac cggttagcaa ttgttttttc gatgagtttg gacaaaccac
     2521 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt
     2581 tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt
     2641 tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct acaaatgtgg
     2701 tacttaagag ggggagacca aagggcgaga cgttaaggcc tcacgtgaca tgtgagcaaa
     2761 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     2821 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     2881 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     2941 gaccctgccg cttacgggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     3001 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     3061 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     3121 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     3181 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     3241 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     3301 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     3361 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     3421 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgccgtctca
     3481 gaagaactcg tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag cggcgatacc
     3541 gtaaagcacg aggaagcggt cagcccattc gccgccaagc tcttcagcaa tatcacgggt
     3601 agccaacgct atgtcctgat agcggtccgc cacacccagc cggccacagt cgatgaatcc
     3661 agaaaagcgg ccattttcca ccatgatatt cggcaagcag gcatcgccat gggtcacgac
     3721 gagatcctcg ccgtcgggca tgctcgcctt gagcctggcg aacagttcgg ctggcgcgag
     3781 cccctgatgc tcttcgtcca gatcatcctg atcgacaaga ccggcttcca tccgagtacg
     3841 tgctctctcg atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg gatcaagcgt
     3901 atgcagccgc cgcattgcat cagccatgat ggatactttc tcggcaggag caaggtgaga
     3961 tgacaggaga tcctgccccg gcacttcgcc caatagcagc cagtcccttc ccgcttcagt
     4021 gacaacgtcg agtacagctg cgcaaggaac gcccgtcgtg gccagccacg atagccgcgc
     4081 tgcctcgtct tgcagttcat tcagggcacc ggacaggtcg gtcttgacaa aaagaaccgg
     4141 gcgcccctgc gctgacagcc ggaacacggc ggcatcagag cagccgattg tctgttgtgc
     4201 ccagtcatag ccgaatagcc tctccaccca agcggccgga gaacctgcgt gcaatccatc
     4261 ttgttcaatc ataatattat tgaagcattt atcagggttc gtctcgtccc ggtctcctcc
     4321 catgca
//
