LOCUS       sequence-125551-        5254 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient expression of humanized MS2 (V75E/A81G), fused to mCherry 
            in mammalian cells, under a UBC promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5254)
  AUTHORS   Shechner DM, Hacisuleyman E, Younger ST, Rinn JL
  TITLE     Multiplexable, locus-specific targeting of long RNAs with 
            CRISPR-Display.
  JOURNAL   Nat Methods. 2015 Jul;12(7):664-70. doi: 10.1038/nmeth.3433. Epub 
            2015 Jun 1.
  PUBMED    26030444
REFERENCE   2  (bases 1 to 5254)
  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"; doi: 
            "10.1038/nmeth.3433"; journalName: "Nat Methods"; date: "2015-07"; 
            volume: "12"; issue: "7"; pages: "664-70"
COMMENT     Sequence Label: pUBC_V5_MS2-mCherry
FEATURES             Location/Qualifiers
     source          1..5254
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(1..792,793..861))
                     /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     624..643
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(862..966)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1034..1052
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1084..1106)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1200..1219
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1413..1435
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1427..1444
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1428..1444
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1468..2679
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     2516..2535
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     regulatory      2697..2706
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2709..2729
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2736..2777
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             3207..3917
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGREFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(3352..3370)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3620..3639
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     3814..3837
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(3964..3983)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4018..4037
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4094..4110)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4094..4110)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4107..4129)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4118..4134
                     /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(4142..4148,4149..4166,4167..4172))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4187..4208
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4325..4342)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4496..5084)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4576..4595)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
       61 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
      121 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
      181 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
      241 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
      301 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
      361 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
      421 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
      481 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
      541 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
      601 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
      661 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
      721 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
      781 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
      841 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
      901 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
      961 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
     1021 attaacctat aaaaataggc gtatcacgag gccctttgcg cgtttcggtg atgacggtga
     1081 aaacctctga cacatgcagc tcccggtcac agcttgtctg taagcggatg ccgggagcag
     1141 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc ttaactatgc
     1201 ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg
     1261 cgtaaggaga aaataccgca tcaggcgcca ttcgccattc aggctgcgca actgttggga
     1321 agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc
     1381 aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc
     1441 cagtgaattc gagctcggta cacgtatgtc gagggcgccg ggttttggcg cctcccgcgg
     1501 gcgcccccct cctcacggcg agcgctgcca cgtcagacga agggcgcagc gagcgtcctg
     1561 atccttccgc ccggacgctc aggacagcgg cccgctgctc ataagactcg gccttagaac
     1621 cccagtatca gcagaaggac attttaggac gggacttggg tgactctagg gcactggttt
     1681 tctttccaga gagcggaaca ggcgaggaaa agtagtccct tctcggcgat tctgcggagg
     1741 gatctccgtg gggcggtgaa cgccgatgat tatataagga cgcgccgggt gtggcacagc
     1801 tagttccgtc gcagccggga tttgggtcgc ggttcttgtt tgtggatcgc tgtgatcgtc
     1861 acttggtgag tagcgggctg ctgggctggc cggggctttc gtggccgccg ggccgctcgg
     1921 tgggacggaa gcgtgtggag agaccgccaa gggctgtagt ctgggtccgc gagcaaggtt
     1981 gccctgaact gggggttggg gggagcgcag caaaatggcg gctgttcccg agtcttgaat
     2041 ggaagacgct tgtgaggcgg gctgtgaggt cgttgaaaca aggtgggggg catggtgggc
     2101 ggcaagaacc caaggtcttg aggccttcgc taatgcggga aagctcttat tcgggtgaga
     2161 tgggctgggg caccatctgg ggaccctgac gtgaagtttg tcactgactg gagaactcgg
     2221 tttgtcgtct gttgcggggg cggcagttat ggcggtgccg ttgggcagtg cacccgtacc
     2281 tttgggagcg cgcgccctcg tcgtgtcgtg acgtcacccg ttctgttggc ttataatgca
     2341 gggtggggcc acctgccggt aggtgtgcgg taggcttttc tccgtcgcag gacgcagggt
     2401 tcgggcctag ggtaggctct cctgaatcga caggcgccgg acctctggtg aggggaggga
     2461 taagtgaggc gtcagtttct ttggtcggtt ttatgtacct atcttcttaa gtagctgaag
     2521 ctccggtttt gaactatgcg ctcggggttg gcgagtgtgt tttgtgaagt tttttaggca
     2581 ccttttgaaa tgtaatcatt tgggtcaata tgtaattttc agtgttagac tagtaaattg
     2641 tccgctaaat tctggccgtt tttggctttt ttgttagacc tggaggtacc gccaccgcca
     2701 ccatggctcc aaagaagaag cgtaaggtag gcagcggcaa gcccatcccc aaccccctgc
     2761 tgggcctgga cagcaccggc agcgctagca atttcactca atttgtgctc gtggataacg
     2821 gaggaacggg cgacgtgacc gtggcaccaa gcaactttgc taatggagtg gccgagtgga
     2881 ttagctcaaa tagcaggtcc caggcttaca aggtcacctg tagtgtccgg caatcatccg
     2941 cccagaatcg gaaatatact attaaggtag aagtgccgaa agtggcaact caaaccgtgg
     3001 ggggcgagga attgcctgtg gccggctgga gaagctacct caacatggaa ctcaccattc
     3061 caatcttcgc cacaaacagc gactgtgagc tcatcgtcaa agccatgcag ggtctgctga
     3121 aggacggcaa tcctattcca tctgcaatcg cggccaattc cggcatttat ggatccaagg
     3181 gctcgggctc gacctcgggc tccggaatgg tgagcaaggg cgaggaggac aacatggcca
     3241 tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg ctccgtgaac ggccgcgagt
     3301 tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc gccaagctga
     3361 aggtgaccaa gggcggcccc ctgcccttcg cctgggacat cctgtcccct cagttcatgt
     3421 acggctccaa ggcctacgtg aagcaccccg ccgacatccc cgactacttg aagctgtcct
     3481 tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg
     3541 tgacccagga ctcctccctg caggacggcg agttcatcta caaggtgaag ctgcgcggca
     3601 ccaacttccc ctccgacggc cccgtaatgc agaagaagac catgggctgg gaggcctcct
     3661 ccgagcggat gtaccccgag gacggcgccc tgaagggcga gatcaagcag aggctgaagc
     3721 tgaaggacgg cggccactac gacgccgagg tcaagaccac ctacaaggcc aagaagcccg
     3781 tgcagctgcc cggcgcctac aacgtcaaca tcaagctgga catcacctcc cacaacgagg
     3841 actacaccat cgtggaacag tacgagcgcg ccgagggccg ccactccacc ggcggcatgg
     3901 acgagctgta caagtaatga taagtcgact tgtttattgc agcttataat ggttacaaat
     3961 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     4021 gtttgtccaa actcatcaat gtatcttatt cgactgttta aacctgcagg catgcaagct
     4081 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     4141 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     4201 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     4261 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     4321 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     4381 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     4441 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4501 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     4561 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     4621 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     4681 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     4741 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     4801 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     4861 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     4921 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     4981 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5041 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5101 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5161 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5221 tctaaagtat atatgagtaa acttggtctg acag
//
