LOCUS       sequence-124596-        4586 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient expression of humanized A. fulgidus L7Ae, fused to the 
            VP64 transcription activator, in mammalian cells, under a UBC 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4586)
  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 4586)
  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_HA_L7Ae-VP64
FEATURES             Location/Qualifiers
     source          1..4586
                     /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     1056..1075
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1269..1291
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1283..1300
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1284..1300
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2359..2378
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     regulatory      2540..2549
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2552..2572
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2576..2596
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2600..2620
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2624..2650
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             2624..2650
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3062..3211
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     primer_bind     complement(3297..3316)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3351..3370
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3426..3442)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3426..3442)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3439..3461)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3450..3466
                     /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(3474..3480,3481..3498,3499..3504))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3519..3540
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3657..3674)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3828..4416)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3908..3927)
                     /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 ctatgcggca tcagagcaga ttgtactgag
     1081 agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
     1141 gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
     1201 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
     1261 agggttttcc cagtcacgac gttgtaaaac gacggccagt gaattcgagc tcggtacgcg
     1321 ccgggttttg gcgcctcccg cgggcgcccc cctcctcacg gcgagcgctg ccacgtcaga
     1381 cgaagggcgc agcgagcgtc ctgatccttc cgcccggacg ctcaggacag cggcccgctg
     1441 ctcataagac tcggccttag aaccccagta tcagcagaag gacattttag gacgggactt
     1501 gggtgactct agggcactgg ttttctttcc agagagcgga acaggcgagg aaaagtagtc
     1561 ccttctcggc gattctgcgg agggatctcc gtggggcggt gaacgccgat gattatataa
     1621 ggacgcgccg ggtgtggcac agctagttcc gtcgcagccg ggatttgggt cgcggttctt
     1681 gtttgtggat cgctgtgatc gtcacttggt gagtagcggg ctgctgggct ggccggggct
     1741 ttcgtggccg ccgggccgct cggtgggacg gaagcgtgtg gagagaccgc caagggctgt
     1801 agtctgggtc cgcgagcaag gttgccctga actgggggtt ggggggagcg cagcaaaatg
     1861 gcggctgttc ccgagtcttg aatggaagac gcttgtgagg cgggctgtga ggtcgttgaa
     1921 acaaggtggg gggcatggtg ggcggcaaga acccaaggtc ttgaggcctt cgctaatgcg
     1981 ggaaagctct tattcgggtg agatgggctg gggcaccatc tggggaccct gacgtgaagt
     2041 ttgtcactga ctggagaact cggtttgtcg tctgttgcgg gggcggcagt tatggcggtg
     2101 ccgttgggca gtgcacccgt acctttggga gcgcgcgccc tcgtcgtgtc gtgacgtcac
     2161 ccgttctgtt ggcttataat gcagggtggg gccacctgcc ggtaggtgtg cggtaggctt
     2221 ttctccgtcg caggacgcag ggttcgggcc tagggtaggc tctcctgaat cgacaggcgc
     2281 cggacctctg gtgaggggag ggataagtga ggcgtcagtt tctttggtcg gttttatgta
     2341 cctatcttct taagtagctg aagctccggt tttgaactat gcgctcgggg ttggcgagtg
     2401 tgttttgtga agttttttag gcaccttttg aaatgtaatc atttgggtca atatgtaatt
     2461 ttcagtgtta gactagtaaa ttgtccgcta aattctggcc gtttttggct tttttgttag
     2521 acctggaggt acggccaccg ccaccatgga ccctaaaaaa aagagaaagg tggacccaaa
     2581 gaagaaacgc aaagttgatc cgaagaagaa aaggaaagtt ggctacccct acgacgtgcc
     2641 cgactacgcc ggaagtggtg ccggcaccgg cggcatgtac gtgcgcttcg aggtgcccga
     2701 ggacatgcag aacgaggccc tgagcctgct ggaaaaagtg cgcgagagcg gcaaagtgaa
     2761 gaagggcacc aacgaaacca ccaaggccgt ggaacggggc ctggccaagc tggtgtatat
     2821 cgccgaggac gtggaccccc ccgagattgt ggcccatctg cccctgctgt gcgaagagaa
     2881 gaacgtgccc tacatctacg tgaagtccaa gaacgacctg ggcagagccg tgggcatcga
     2941 ggtgccatgt gcctctgccg ccatcatcaa cgagggcgag ctgcggaaag aactgggcag
     3001 cctggtggaa aagatcaagg gcctgcagaa gggttccggt ggatccggtt ccggacgggc
     3061 tgacgcattg gacgattttg atctggatat gctgggaagt gacgccctcg atgattttga
     3121 ccttgacatg cttggttcgg atgcccttga tgactttgac ctcgacatgc tcggcagtga
     3181 cgcccttgat gatttcgacc tggacatgct gattaactga ggtacctgaa taaatgaggg
     3241 cccgtttaaa cccgctgatc agttgtttat tgcagcttat aatggttaca aataaagcaa
     3301 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     3361 caaactcatc aatgtatctt atcgactgtt taaacctgca ggcatgcaag cttggcgtaa
     3421 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     3481 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     3541 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     3601 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     3661 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     3721 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     3781 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     3841 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     3901 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     3961 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     4021 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     4081 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     4141 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     4201 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     4261 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     4321 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     4381 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     4441 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     4501 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     4561 atatatgagt aaacttggtc tgacag
//
