LOCUS       sequence-232164-        7441 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CRISPR-Sirius plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7441)
  AUTHORS   Ma H, Tu LC, Naseri A, Chung YC, Grunwald D, Zhang S, Pederson T
  TITLE     CRISPR-Sirius: RNA scaffolds for signal amplification in genome 
            imaging.
  JOURNAL   Nat Methods. 2018 Nov;15(11):928-931. doi: 
            10.1038/s41592-018-0174-0. Epub 2018 Oct 30.
  PUBMED    30377374
REFERENCE   2  (bases 1 to 7441)
  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/s41592-018-0174-0"; journalName: "Nat Methods"; date: 
            "2018-11"; volume: "15"; issue: "11"; pages: "928-931"
COMMENT     Sequence Label: pHAGE-EFS-MCP-HALOnls
FEATURES             Location/Qualifiers
     source          1..7441
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..634
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1721..1765
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2058..2175
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2265..2476
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2522..2531
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2906..3793
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="AEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSY
                     VWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIH
                     DWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIID
                     QNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVE
                     EYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLI
                     GSEIARWLSTLEISG"
     CDS             3800..3820
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_feature    3831..4419
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3884..3904)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4302..4313)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4494..4727
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(4846..4864)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4932..5036
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5037..5105,5106..5897)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5255..5274)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6068..6656
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6557..6576
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6810..6827
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7067..7088
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7103..7108,7109..7126,7127..7133)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7141..7157
                     /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)."
     primer_bind     7146..7168
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7165..7181
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7165..7181
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7231..7250
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccgaa ttgacaaatg
     2041 gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa
     2101 agaatagtag acataatagc aacagacata caaactaaag aattacaaaa acaaattaca
     2161 aaaattcaaa attttcgggt ttattacagg gacagcagag atccagtttg gactagcgat
     2221 gctagctgat gcgggccctt cgagtggctc cggtgcccgt cagtgggcag agcgcacatc
     2281 gcccacagtc cccgagaagt tggggggagg ggtcggcaat tgaaccggtg cctagagaag
     2341 gtggcgcggg gtaaactggg aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg
     2401 tgggggagaa ccgtatataa gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt
     2461 tgccgccaga acacaggtgt cgtgacgcgg ttccggttaa ggtgaacgcg tgcaggctgg
     2521 cgccaccatg gcttctaact ttactcagtt cgttctcgtc gacaatggcg gaactggcga
     2581 cgtgactgtc gccccaagca acttcgctaa cgggatcgct gaatggatca gctctaactc
     2641 gcgttcacag gcttacaaag taacctgtag cgttcgtcag agctctgcgc agaatcgcaa
     2701 atacaccatc aaagtcgagg tgcctaaagg cgcctggcgt tcgtacttaa atatggaact
     2761 aaccattcca attttcgcca cgaattccga ctgcgagctt attgttaagg caatgcaagg
     2821 tctcctaaaa gatggaaacc cgattccctc agcaatcgca gcaaactccg gcatctacgg
     2881 gggatctggt agcggcgctg gatccgcaga aatcggtact ggctttccat tcgaccccca
     2941 ttatgtggaa gtcctgggcg agcgcatgca ctacgtcgat gttggtccgc gcgatggcac
     3001 ccctgtgctg ttcctgcacg gtaacccgac ctcctcctac gtgtggcgca acatcatccc
     3061 gcatgttgca ccgacccatc gctgcattgc tccagacctg atcggtatgg gcaaatccga
     3121 caaaccagac ctgggttatt tcttcgacga ccacgtccgc ttcatggatg ccttcatcga
     3181 agccctgggt ctggaagagg tcgtcctggt cattcacgac tggggctccg ctctgggttt
     3241 ccactgggcc aagcgcaatc cagagcgcgt caaaggtatt gcatttatgg agttcatccg
     3301 ccctatcccg acctgggacg aatggccaga atttgcccgc gagaccttcc aggccttccg
     3361 caccaccgac gtcggccgca agctgatcat cgatcagaac gtttttatcg agggtacgct
     3421 gccgatgggt gtcgtccgcc cgctgactga agtcgagatg gaccattacc gcgagccgtt
     3481 cctgaatcct gttgaccgcg agccactgtg gcgcttccca aacgagctgc caatcgccgg
     3541 tgagccagcg aacatcgtcg cgctggtcga agaatacatg gactggctgc accagtcccc
     3601 tgtcccgaag ctgctgttct ggggcacccc aggcgttctg atcccaccgg ccgaagccgc
     3661 tcgcctggcc aaaagcctgc ctaactgcaa ggctgtggac atcggcccgg gtctgaatct
     3721 gctgcaagaa gacaacccgg acctgatcgg cagcgagatc gcgcgctggc tgtcgacgct
     3781 ggagatttcc ggcctcgagc caaagaaaaa gcggaaagtg taatctagat aatcaacctc
     3841 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
     3901 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
     3961 ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg
     4021 tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca
     4081 ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg
     4141 cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg
     4201 acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg
     4261 ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg
     4321 accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc
     4381 ctcagacgag tcggatctcc ctttgggccg cctccccgcc tgagatcctt taagaccaat
     4441 gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg
     4501 gctaattcac tcccaacgaa gacaagatct gctttttgct tgtactgggt ctctctggtt
     4561 agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc ttaagcctca
     4621 ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa
     4681 ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcagta gtagttcatg
     4741 tcatcttatt attcagtatt tataacttgc aaagaaatga atatcagaga gtgagaggcc
     4801 cgggttaatt aaggaaaggg ctagatcatt cttgaagacg aaagggcctc gtgatacgcc
     4861 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     4921 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     4981 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     5041 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     5101 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     5161 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     5221 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg
     5281 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     5341 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     5401 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     5461 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     5521 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     5581 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     5641 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     5701 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     5761 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     5821 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     5881 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     5941 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     6001 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     6061 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     6121 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     6181 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     6241 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     6301 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     6361 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     6421 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     6481 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     6541 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     6601 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     6661 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     6721 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     6781 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     6841 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agcaagctca
     6901 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt
     6961 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tccccgtggc
     7021 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
     7081 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
     7141 ttgtgagcgg ataacaattt cacacaggaa acagctatga catgattacg aatttcacaa
     7201 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     7261 atcatgtctg gatcaactgg ataactcaag ctaaccaaaa tcatcccaaa cttcccaccc
     7321 cataccctat taccactgcc aattacctgt ggtttcattt actctaaacc tgtgattcct
     7381 ctgaattatt ttcattttaa agaaattgta tttgttaaat atgtactaca aacttagtag
     7441 t
//
