LOCUS       sequence-239522-        6119 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses 5S rRNA fused to two dimeric Broccoli aptamers in the F30 
            scaffold in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6119)
  AUTHORS   Filonov GS, Jaffrey SR
  TITLE     RNA Imaging with Dimeric Broccoli in Live Bacterial and Mammalian 
            Cells.
  JOURNAL   Curr Protoc Chem Biol. 2016 Mar 16;8(1):1-28. doi: 
            10.1002/9780470559277.ch150174.
  PUBMED    26995352
REFERENCE   2  (bases 1 to 6119)
  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: 
            "10.1002/9780470559277"; journalName: "Curr Protoc Chem Biol"; date:
            "2016-03-16- 16"; volume: "8"; issue: "1"; pages: "1-28"
COMMENT     Sequence Label: pAV5S-F30-2xdBroccoli
FEATURES             Location/Qualifiers
     source          1..6119
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    37..171
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(596..616)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     intron          746..811
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             complement(1231..2025)
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1342..1361)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1952..1971
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(2382..2711)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2396..2531
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2696..2716
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    3284..3405
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     repeat_region   3454..3583
                     /label=AAV2 ITR
     rep_origin      complement(3677..4132)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3814..3835)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4026..4045
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4159..4263
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4264..4332,4333..5124)
                     /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     complement(4482..4501)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5295..5883
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5784..5803
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gtgaattacg tcatagggtt agggaggtcc tggatcgatc cagacatgat aagatacatt
       61 gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt
      121 tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac
      181 aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag
      241 taaaacctct acaaatgtgg tatggctgat tatgatctct agtcaaggca ctatacatca
      301 aatattcctt attaacccct ttacaaatta aaaagctaaa ggtacacaat ttttgagcat
      361 agttattaat agcagacact ctatgcctgt gtggagtaag aaaaaacagt atgttatgat
      421 tataactgtt atgcctactt ataaaggtta cagaatattt ttccataatt ttcttgtata
      481 gcagtgcagc tttttccttt gtggtgtaaa tagcaaagca agcaagagtt ctattactaa
      541 acacagcatg actcaaaaaa cttagcaatt ctgaaggaaa gtccttgggg tcttctacct
      601 ttctcttctt ttttggagga gtagaatgtt gagagtcagc agtagcctca tcatcactag
      661 atggcatttc ttctgagcaa aacaggtttt cctcattaaa ggcattccac cactgctccc
      721 attcatcagt tccataggtt ggaatctaaa atacacaaac aattagaatc agtagtttaa
      781 cacattatac acttaaaaat tttatattta ccttagagct ttaaatctct gtaggtagtt
      841 tgtccaatta tgtcacacca cagaagtaag gttccttcac aaagatccgg accaaagcgg
      901 ccatcgtgcc tccccactcc tgcagttcgg gggcatggat gcgcggatag ccgctgctgg
      961 tttcctggat gccgacggat ttgcactgcc ggtagaactc cgcgaggtcg tccagcctca
     1021 ggcagcagct gaaccaactc gcgaggggat cgagcccggg gtgggcgaag aactccagca
     1081 tgagatcccc gcgctggagg atcatccagc cggcgtcccg gaaaacgatt ccgaagccca
     1141 acctttcata gaaggcggcg gtggaatcga aatctcgtga tggcaggttg ggcgtcgctt
     1201 ggtcggtcat ttcgaacccc agagtcccgc tcagaagaac tcgtcaagaa ggcgatagaa
     1261 ggcgatgcgc tgcgaatcgg gagcggcgat accgtaaagc acgaggaagc ggtcagccca
     1321 ttcgccgcca agctcttcag caatatcacg ggtagccaac gctatgtcct gatagcggtc
     1381 cgccacaccc agccggccac agtcgatgaa tccagaaaag cggccatttt ccaccatgat
     1441 attcggcaag caggcatcgc catgggtcac gacgagatcc tcgccgtcgg gcatgcgcgc
     1501 cttgagcctg gcgaacagtt cggctggcgc gagcccctga tgctcttcgt ccagatcatc
     1561 ctgatcgaca agaccggctt ccatccgagt acgtgctcgc tcgatgcgat gtttcgcttg
     1621 gtggtcgaat gggcaggtag ccggatcaag cgtatgcagc cgccgcattg catcagccat
     1681 gatggatact ttctcggcag gagcaaggtg agatgacagg agatcctgcc ccggcacttc
     1741 gcccaatagc agccagtccc ttcccgcttc agtgacaacg tcgagcacag ctgcgcaagg
     1801 aacgcccgtc gtggccagcc acgatagccg cgctgcctcg tcctgcagtt cattcagggc
     1861 accggacagg tcggtcttga caaaaagaac cgggcgcccc tgcgctgaca gccggaacac
     1921 ggcggcatca gagcagccga ttgtctgttg tgcccagtca tagccgaata gcctctccac
     1981 ccaagcggcc ggagaacctg cgtgcaatcc atcttgttca atcatgcgaa acgatcctca
     2041 tcctgtctct tgatcagatc ttgatcccct gcgccatcag atccttggcg gcaagaaagc
     2101 catccagttt actttgcagg gcttcccaac cttaccagag ggcgccccag ctggcaattc
     2161 cggttcgctt gctgtccata aaaccgccca gtctagctat cgccatgtaa gcccactgca
     2221 agctacctgc tttctctttg cgcttgcgtt ttcccttgtc cagatagccc agtagctgac
     2281 attcatccgg ggtcagcacc gtttctgcgg actggctttc tacgtgttcc gcttccttta
     2341 gcagcccttg cgccctgagt gcttgcggca gcgtgaagct ttttgcaaaa gcctaggcct
     2401 ccaaaaaagc ctcctcacta cttctggaat agctcagagg ccgaggcggc ctcggcctct
     2461 gcataaataa aaaaaattag tcagccatgg ggcggagaat gggcggaact gggcggagtt
     2521 aggggcggga tgggcggagt taggggcggg actatggttg ctgactaatt gagatgcatg
     2581 ctttgcatac ttctgcctgc tggggagcct ggggactttc cacacctggt tgctgactaa
     2641 ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt tccacaccct
     2701 aactgacaca cattccacag ccggatccgg ccccgggctg gcggtgtcgg atgcaatccg
     2761 gcgggcacgg ccgggccggg ctgggctctt ggggcagcca ggcgcctcct tcagcgtcta
     2821 cggccatacc accctgaacg cgcccgatct cgtctgatct cggaagctaa gcagggtcgg
     2881 gcctggttag tacttggatg ggagaccgcc tgggaatacc gggtgctgta ggcgtcgact
     2941 tgccatgtgt atgtgggaga cggtcgggtc catctgagac ggtcgggtcc agatattcgt
     3001 atctgtcgag tagagtgtgg gctcagatgt cgagtagagt gtgggctccc acatactctg
     3061 atgatccaga cggtcgggtc catctgagac ggtcgggtcc agatattcgt atctgtcgag
     3121 tagagtgtgg gctcagatgt cgagtagagt gtgggctgga tcattcatgg caatctagag
     3181 cggacttcgg tccgcttttt actaggacct gcaggcatgc aagcttgacg tcggttaccg
     3241 atatccatat ggcggccgca tcgatctcga gccgcggact agtaacttgt ttattgcagc
     3301 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     3361 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tcttacgtag
     3421 ataagtagca tggcgggtta atcattaact acaaggaacc cctagtgatg gagttggcca
     3481 ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc
     3541 cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagagaggga cagatctgca
     3601 gatctgctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag
     3661 cctgaatggc gaatggaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt
     3721 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca
     3781 aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta
     3841 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
     3901 cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
     3961 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
     4021 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg
     4081 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc
     4141 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     4201 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     4261 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     4321 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     4381 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     4441 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     4501 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     4561 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     4621 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     4681 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     4741 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     4801 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     4861 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     4921 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     4981 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     5041 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     5101 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     5161 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     5221 atgaccaaaa tcccttaacg taagttttcg ttccactgag cgtcagaccc cgtagaaaag
     5281 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     5341 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     5401 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     5461 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     5521 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     5581 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     5641 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     5701 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     5761 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     5821 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     5881 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     5941 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     6001 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgagcgagcg
     6061 cgcagagagg gagtggccaa ctccatcact aggggttcct ggaggggtgg agtcgtgac
//
