LOCUS       sequence-431684-        4342 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses BxbI-BP-NLS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4342)
  AUTHORS   Autio MI, Motakis E, Perrin A, Bin Amin T, Tiang Z, Do DV, Wang J, 
            Tan JKM, Ding SSL, Tan WX, Lee CJM, Teo AKK, Foo R
  TITLE     Computationally defined and in vitro validated putative genomic safe
            harbour loci for transgene expression in human cells.
  JOURNAL   Elife. 2024 Jan 2;13:e79592. doi: 10.7554/eLife.79592.
  PUBMED    38164941
REFERENCE   2  (bases 1 to 4342)
  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: "Elife."; 
            date: "2024-01-2"; volume: "13:e79592. doi"; pages: " 
            10.7554/eLife.79592"
COMMENT     Sequence Label: pMIA22
FEATURES             Location/Qualifiers
     source          1..4342
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        77..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          676..808
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     CDS             2348..2368
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(2378..2397)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    2407..2488
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     2432..2451
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(2686..3274)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2766..2785)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3355..4149)
                     /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(3355..4149)
                     /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(3466..3485)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4076..4095
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 ggcattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag aggtcgttta gtgaaccgtc
      601 agatcactag tagctttatt gcggtagttt atcacagtta aattgctaac gcagtcagtg
      661 ctcgactgat cacaggtaag tatcaaggtt acaagacagg tttaaggagg ccaatagaaa
      721 ctgggcttgt cgagacagag aagattcttg cgtttctgat aggcacctat tggtcttact
      781 gacatccact ttgcctttct ctccacaggc caccatgcgg gcactcgttg taatcaggtt
      841 gtctcgagtt acggatgcga ccacgtcccc ggagagacaa ctcgaatcat gccagcagct
      901 ttgtgctcag agaggatggg acgttgtggg cgtggccgaa gatctcgacg tatcaggggc
      961 ggtggacccc ttcgatagaa agagaagacc aaatcttgcc aggtggctgg ctttcgaaga
     1021 gcaacctttt gacgtaatcg tggcgtatag agtagacagg ctcacgaggt ctatccggca
     1081 ccttcaacaa ttggttcact gggcagaaga ccacaagaag ttggtggttt cagcaactga
     1141 ggcgcacttc gatactacca ctccatttgc ggcagtcgtg attgccctta tgggtacggt
     1201 tgcccaaatg gaactggagg ctattaagga gcggaatagg agtgccgcac attttaatat
     1261 ccgggctggc aaatatcgcg gatcactccc cccgtggggt tacctcccta cccgcgtaga
     1321 cggtgagtgg aggctcgtac cggaccccgt tcagagagag cggatcttgg aagtgtatca
     1381 cagagtagtg gacaaccatg aacctttgca tctcgtagcg catgatctca atcgcagggg
     1441 ggtactctct ccaaaagatt atttcgcgca acttcagggt cgggagcccc agggaagaga
     1501 atggtctgca acagctctca agcggagtat gataagtgaa gccatgcttg gatacgccac
     1561 tcttaacggg aaaaccgtca gagacgacga tggcgcacca ctcgtcaggg cggagcccat
     1621 tttgacacgg gaacaacttg aggcgcttcg ggcagaactg gttaagacca gtcgcgctaa
     1681 gcctgcagtg tctactccct ccctcttgct cagagtcctg ttttgcgcag tttgtggtga
     1741 gcccgcctat aaatttgctg gaggcggacg gaagcaccca agataccgct gcagaagcat
     1801 gggatttccc aaacactgcg ggaatggaac agtcgctatg gccgagtggg acgccttttg
     1861 cgaagagcag gtgcttgatt tgctgggaga cgctgaacgg ctggaaaaag tgtgggttgc
     1921 agggtcagat agtgccgtag agttggcgga ggttaacgca gaacttgtgg atctgacttc
     1981 tttgattggc tcccccgcgt atcgggccgg aagcccccaa cgagaagccc ttgacgcccg
     2041 aattgcggcc ctcgcagccc gacaagagga actggaagga cttgaagccc gccctagtgg
     2101 atgggagtgg cgggagacag gacaaaggtt tggagactgg tggagagaac aagatactgc
     2161 ggcgaagaac acatggctgc gatcaatgaa cgtccggctg acgtttgacg ttcggggcgg
     2221 cttgacaagg actattgatt tcggggatct gcaagaatat gaacagcatc ttagactggg
     2281 ctctgtagtt gaaaggcttc acaccggtat gtccaaaagg accgcagatg gctccgagtt
     2341 tgagtcaccg aagaagaaga ggaaggtcga ataataggat gagtttggac aaaccacaac
     2401 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     2461 aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca
     2521 ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtac
     2581 ttaagagggg gagaccaaag ggcgagacgt taaggcctca cgtgacatgt gagcaaaagg
     2641 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     2701 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     2761 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     2821 cctgccgctt acgggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     2881 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     2941 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     3001 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     3061 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     3121 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     3181 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     3241 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     3301 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatgc cgtctcagaa
     3361 gaactcgtca agaaggcgat agaaggcgat gcgctgcgaa tcgggagcgg cgataccgta
     3421 aagcacgagg aagcggtcag cccattcgcc gccaagctct tcagcaatat cacgggtagc
     3481 caacgctatg tcctgatagc ggtccgccac acccagccgg ccacagtcga tgaatccaga
     3541 aaagcggcca ttttccacca tgatattcgg caagcaggca tcgccatggg tcacgacgag
     3601 atcctcgccg tcgggcatgc tcgccttgag cctggcgaac agttcggctg gcgcgagccc
     3661 ctgatgctct tcgtccagat catcctgatc gacaagaccg gcttccatcc gagtacgtgc
     3721 tctctcgatg cgatgtttcg cttggtggtc gaatgggcag gtagccggat caagcgtatg
     3781 cagccgccgc attgcatcag ccatgatgga tactttctcg gcaggagcaa ggtgagatga
     3841 caggagatcc tgccccggca cttcgcccaa tagcagccag tcccttcccg cttcagtgac
     3901 aacgtcgagt acagctgcgc aaggaacgcc cgtcgtggcc agccacgata gccgcgctgc
     3961 ctcgtcttgc agttcattca gggcaccgga caggtcggtc ttgacaaaaa gaaccgggcg
     4021 cccctgcgct gacagccgga acacggcggc atcagagcag ccgattgtct gttgtgccca
     4081 gtcatagccg aatagcctct ccacccaagc ggccggagaa cctgcgtgca atccatcttg
     4141 ttcaatcata atattattga agcatttatc agggttcgtc tcgtcccggt ctcctcccat
     4201 gcatgtcaat attggccatt agccatatta ttcattggtt atatagcata aatcaatatt
     4261 ggctattggc cattgcatac gttgtatcta tatcataata tgtacattta tattggctca
     4321 tgtccaatat gaccgccatg tt
//
