LOCUS       sequence-239367-        2800 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MS2-NLS (no stop codon) with GreenGate C-D flanking sequences.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2800)
  AUTHORS   Lee JE, Neumann M, Duro DI, Schmid M
  TITLE     CRISPR-based tools for targeted transcriptional and epigenetic 
            regulation in plants.
  JOURNAL   PLoS One. 2019 Sep 26;14(9):e0222778. doi: 
            10.1371/journal.pone.0222778. eCollection 2019.
  PUBMED    31557222
REFERENCE   2  (bases 1 to 2800)
  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: "PLoS One.";
            date: "2019-09-26"; volume: "14(9):e0222778. doi"; pages: " 
            10.1371/journal.pone.0222778. eCollection 2019"
COMMENT     Sequence Label: pMH186
FEATURES             Location/Qualifiers
     source          1..2800
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     46..68
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     60..77
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     61..77
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     88..107
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        88..106
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             111..122
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             164..550
                     /codon_start=1
                     /product="bacteriophage MS2 coat protein"
                     /label=MS2-N55K
                     /note="binds with high affinity to a specific stem-loop 
                     structure in the viral RNA (Lim et al., 1994)"
                     /translation="ASNFTQFVLVDNGGTGDVTVAPSNFANGVAEWISSNSRSQAYKVT
                     CSVRQSSAQKRKYTIKVEVPKVATQTVGGVELPVAAWRSYLNMELTIPIFATNSDCELI
                     VKAMQGLLKDGNPIPSAIAANSGIY"
     CDS             605..625
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(774..791)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(884..1472)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(964..983)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1646..2440)
                     /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"
     promoter        complement(2441..2545)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
       61 gtaaaacgac ggccagtgag cgcgacgtaa tacgactcac tatagggcga attgaaggaa
      121 ggccgtcaag gccgcatagt gaagcttggt ctcaggctcc atggcttcta attttactca
      181 atttgttttg gttgataatg gaggaactgg agatgttact gttgctcctt ctaattttgc
      241 taatggagtt gctgaatgga tttcttctaa ttcaagatct caggcttata aggttacttg
      301 ttctgttaga caatcttctg ctcaaaagag aaagtatact attaaggttg aagttcctaa
      361 ggttgctact caaactgttg gaggagttga attgcctgtt gctgcttgga gatcttattt
      421 gaatatggaa ttgactattc ctatttttgc tactaattct gattgtgaat tgattgttaa
      481 ggctatgcaa ggattgttga aggatggaaa tcctattcct tctgctattg ctgctaattc
      541 tggaatttat tctgctggag gaggaggatc tggaggagga ggatctggag gaggaggatc
      601 tggacctaag aagaagagaa aggttgctgc tgctggatct tcagtgagac cgaattctcc
      661 tgggcctcat gggccttcct ttcactgccc gctttccagt cgggaaacct gtcgtgccag
      721 ctgcattaac atggtcatag ctgtttcctt gcgtattggg cgctctccgc ttcctcgctc
      781 actgactcgc tgcgctcggt cgttcgggta aagcctgggg tgcctaatga gcaaaaggcc
      841 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
      901 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
      961 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     1021 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     1081 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     1141 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     1201 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     1261 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     1321 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     1381 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     1441 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     1501 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     1561 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     1621 atgagtaaac ttggtctgac agttattaga aaaattcatc cagcagacga taaaacgcaa
     1681 tacgctggct atccggtgcc gcaatgccat acagcaccag aaaacgatcc gcccattcgc
     1741 cgcccagttc ttccgcaata tcacgggtgg ccagcgcaat atcctgataa cgatccgcca
     1801 cgcccagacg gccgcaatca ataaagccgc taaaacggcc attttccacc ataatgttcg
     1861 gcaggcacgc atcaccatgg gtcaccacca gatcttcgcc atccggcatg ctcgctttca
     1921 gacgcgcaaa cagctctgcc ggtgccaggc cctgatgttc ttcatccaga tcatcctgat
     1981 ccaccaggcc cgcttccata cgggtacgcg cacgttcaat acgatgtttc gcctgatgat
     2041 caaacggaca ggtcgccggg tccagggtat gcagacgacg catggcatcc gccataatgc
     2101 tcactttttc tgccggcgcc agatggctag acagcagatc ctgacccggc acttcgccca
     2161 gcagcagcca atcacggccc gcttcggtca ccacatccag caccgccgca cacggaacac
     2221 cggtggtggc cagccagctc agacgcgccg cttcatcctg cagctcgttc agcgcaccgc
     2281 tcagatcggt tttcacaaac agcaccggac gaccctgcgc gctcagacga aacaccgccg
     2341 catcagagca gccaatggtc tgctgcgccc aatcatagcc aaacagacgt tccacccacg
     2401 ctgccgggct acccgcatgc aggccatcct gttcaatcat actcttcctt tttcaatatt
     2461 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     2521 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct aaattgtaag
     2581 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     2641 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     2701 tggccgctac agggcgctcc cattcgccat tcaggctgcg caactgttgg gaagggcgtt
     2761 tcggtgcggg cctcttcgct attacgccag ctggcgaaag
//
