LOCUS       sequence-419977-        7092 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Express NES-dpspCas13b(AAAA)-FseI[ADAR2-DD-splitC].
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7092)
  AUTHORS   Liu Z, Jillette N, Robson P, Cheng AW
  TITLE     Simultaneous multifunctional transcriptome engineering by CRISPR RNA
            scaffold.
  JOURNAL   Nucleic Acids Res. 2023 Aug 11;51(14):e77. doi: 10.1093/nar/gkad547.
  PUBMED    37395412
REFERENCE   2  (bases 1 to 7092)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res."; date: "2023-08-11"; volume: "51(14):e77. doi"; pages: "
            10.1093/nar/gkad547"
COMMENT     Sequence Label: 
            pzk490-pmax-NES-dpspCas13b(AAAA)-FseI[ADAR2-DD-splitC]
FEATURES             Location/Qualifiers
     source          1..7092
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          813..945
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     protein_bind    960..984
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1011..1037
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     protein_bind    complement(5121..5145)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             5198..5239
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(5265..5284)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    5294..5375
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     5319..5338
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(5573..6161)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5653..5672)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6242..7036)
                     /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"
     CDS             complement(6242..7036)
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6353..6372)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     6963..6982
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagg
      721 tcgtttagtg aaccgtcaga tcactagtag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctc gactgatcac aggtaagtat caaggttaca agacaggttt
      841 aaggaggcca atagaaactg ggcttgtcga gacagagaag attcttgcgt ttctgatagg
      901 cacctattgg tcttactgac atccactttg cctttctctc cacagggaaa aaacaattga
      961 caagtttgta caaaaaagca ggctccgaat tcaccggtgc cgccaccatg ctgccaccat
     1021 tggaaaggct tacgctgggc ggaggcggat ctgggcgcgc catgaacatc cccgctctgg
     1081 tggaaaacca gaagaagtac tttggcacct acagcgtgat ggccatgctg aacgctcaga
     1141 ccgtgctgga ccacatccag aaggtggccg atattgaggg cgagcagaac gagaacaacg
     1201 agaatctgtg gtttcacccc gtgatgagcc acctgtacaa cgccaagaac ggctacgaca
     1261 agcagcccga gaaaaccatg ttcatcatcg agcggctgca gagctacttc ccattcctga
     1321 agatcatggc cgagaaccag agagagtaca gcaacggcaa gtacaagcag aaccgcgtgg
     1381 aagtgaacag caacgacatc ttcgaggtgc tgaagcgcgc cttcggcgtg ctgaagatgt
     1441 acagggacct gaccaacgca tacaagacct acgaggaaaa gctgaacgac ggctgcgagt
     1501 tcctgaccag cacagagcaa cctctgagcg gcatgatcaa caactactac acagtggccc
     1561 tgcggaacat gaacgagaga tacggctaca agacagagga cctggccttc atccaggaca
     1621 agcggttcaa gttcgtgaag gacgcctacg gcaagaaaaa gtcccaagtg aataccggat
     1681 tcttcctgag cctgcaggac tacaacggcg acacacagaa gaagctgcac ctgagcggag
     1741 tgggaatcgc cctgctgatc tgcctgttcc tggacaagca gtacatcaac atctttctga
     1801 gcaggctgcc catcttctcc agctacaatg cccagagcga ggaacggcgg atcatcatca
     1861 gatccttcgg catcaacagc atcaagctgc ccaaggaccg gatccacagc gagaagtcca
     1921 acaagagcgt ggccatggat atgctcaacg aagtgaagcg gtgccccgac gagctgttca
     1981 caacactgtc tgccgagaag cagtcccggt tcagaatcat cagcgacgac cacaatgaag
     2041 tgctgatgaa gcggagcagc gacagattcg tgcctctgct gctgcagtat atcgattacg
     2101 gcaagctgtt cgaccacatc aggttccacg tgaacatggg caagctgaga tacctgctgg
     2161 cggccgccgc tacctgcatc gacggccaga ccagagtcag agtgatcgag cagcccctga
     2221 acggcttcgg cagactggaa gaggccgaga caatgcggaa gcaagagaac ggcaccttcg
     2281 gcaacagcgg catccggatc agagacttcg agaacatgaa gcgggacgac gccaatcctg
     2341 ccaactatcc ctacatcgtg gacacctaca cacactacat cctggaaaac aacaaggtcg
     2401 agatgtttat caacgacaaa gaggacagcg ccccactgct gcccgtgatc gaggatgata
     2461 gatacgtggt caagacaatc cccagctgcc ggatgagcac cctggaaatt ccagccatgg
     2521 ccttccacat gtttctgttc ggcagcaaga aaaccgagaa gctgatcgtg gacgtgcaca
     2581 accggtacaa gagactgttc caggccatgc agaaagaaga agtgaccgcc gagaatatcg
     2641 ccagcttcgg aatcgccgag agcgacctgc ctcagaagat cctggatctg atcagcggca
     2701 atgcccacgg caaggatgtg gacgccttca tcagactgac cgtggacgac atgctgaccg
     2761 acaccgagcg gagaatcaag agattcaagg acgaccggaa gtccattcgg agcgccgaca
     2821 acaagatggg aaagagaggc ttcaagcaga tctccacagg caagctggcc gacttcctgg
     2881 ccaaggacat cgtgctgttt cagcccagcg tgaacgatgg cgagaacaag atcaccggcc
     2941 tgaactaccg gatcatgcag agcgccattg ccgtgtacga tagcggcgac gattacgagg
     3001 ccaagcagca gttcaagctg atgttcgaga aggcccggct gatcggcaag ggcacaacag
     3061 agcctcatcc atttctgtac aaggtgttcg cccgcagcat ccccgccaat gccgtcgagt
     3121 tctacgagcg ctacctgatc gagcggaagt tctacctgac cggcctgtcc aacgagatca
     3181 agaaaggcaa cagagtggat gtgcccttca tccggcggga ccagaacaag tggaaaacac
     3241 ccgccatgaa gaccctgggc agaatctaca gcgaggatct gcccgtggaa ctgcccagac
     3301 agatgttcga caatgagatc aagtcccacc tgaagtccct gccacagatg gaaggcatcg
     3361 acttcaacaa tgccaacgtg acctatctga tcgccgagta catgaagaga gtgctggacg
     3421 acgacttcca gaccttctac cagtggaacc gcaactaccg gtacatggac atgcttaagg
     3481 gcgagtacga cagaaagggc tccctgcagc actgcttcac cagcgtggaa gagagagaag
     3541 gcctctggaa agagcgggcc tccagaacag agcggtacag aaagcaggcc agcaacaaga
     3601 tccgcagcaa ccggcagatg agaaacgcca gcagcgaaga gatcgagaca atcctggata
     3661 agcggctgag caacagccgg aacgagtacc agaaaagcga gaaagtgatc cggcgctaca
     3721 gagtgcagga tgccctgctg tttctgctgg ccaaaaagac cctgaccgaa ctggccgatt
     3781 tcgacggcga gaggttcaaa ctgaaagaaa tcatgcccga cgccgagaag ggaatcctga
     3841 gcgagatcat gcccatgagc ttcaccttcg agaaaggcgg caagaagtac accatcacca
     3901 gcgagggcat gaagctgaag aactacggcg acttctttgt gctggctagc gacaagagga
     3961 tcggcaacct gctggaactc gtgggcagcg acatcgtgtc caaagaggat atcatggaag
     4021 agttcaacaa atacgaccag tgcaggcccg agatcagctc catcgtgttc aacctggaaa
     4081 agtgggcctt cgacacatac cccgagctgt ctgccagagt ggaccgggaa gagaaggtgg
     4141 acttcaagag catcctgaaa atcctgctga acaacaagaa catcaacaaa gagcagagcg
     4201 acatcctgcg gaagatccgg aacgccttcg atgcaaacaa ttaccccgac aaaggcgtgg
     4261 tggaaatcaa ggccctgcct gagatcgcca tgagcatcaa gaaggccttt ggggagtacg
     4321 ccatcatgaa gggcggccgc gggggaggcg gttccggtgg cggcggaagc ggaggtggag
     4381 gatcagggcc ggccccagca gatagacacc caaatcgtaa agcaagagga cagctacgga
     4441 ccaaaataga gtctggtcag gggacgattc cagtgcgctc caatgcgagc atccaaacgt
     4501 gggacggggt gctgcaaggg gagcggctgc tcaccatgtc ctgcagtgac aagattgcac
     4561 gctggaacgt ggtgggcatc cagggatcac tgctcagcat tttcgtggag cccatttact
     4621 tctcgagcat catcctgggc agcctttacc acggggacca cctttccagg gccatgtacc
     4681 agcggatctc caacatagag gacctgccac ctctctacac cctcaacaag cctttgctca
     4741 gtggcatcag caatgcagaa gcacggcagc cagggaaggc ccccaacttc agtgtcaact
     4801 ggacggtagg cgactccgct attgaggtca tcaacgccac gactgggaag gatgagctgg
     4861 gccgcgcgtc ccgcctgtgt aagcacgcgt tgtactgtcg ctggatgcgt gtgcacggca
     4921 aggttccctc ccacttacta cgctccaaga ttaccaagcc caacgtgtac catgagtcca
     4981 agctggcggc aaaggagtac caggccgcca aggcgcgtct gttcacagcc ttcatcaagg
     5041 cggggctggg ggcctgggtg gagaagccca ccgagcagga ccagttctca ctcacgaacg
     5101 tttagttaat taagaattcg acccagcttt cttgtacaaa gtggttgata tccagcacag
     5161 tggcggccgc tcgagtctag agggcccgcg gttcgaaggt aagcctatcc ctaaccctct
     5221 cctcggtctc gattctacgc gtaccggtta gcaattgttt tttcgatgag tttggacaaa
     5281 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     5341 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     5401 tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac ctctacaaat
     5461 gtggtactta agagggggag accaaagggc gagacgttaa ggcctcacgt gacatgtgag
     5521 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     5581 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     5641 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     5701 ttccgaccct gccgcttacg ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     5761 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     5821 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     5881 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     5941 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     6001 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     6061 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     6121 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     6181 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgccgt
     6241 ctcagaagaa ctcgtcaaga aggcgataga aggcgatgcg ctgcgaatcg ggagcggcga
     6301 taccgtaaag cacgaggaag cggtcagccc attcgccgcc aagctcttca gcaatatcac
     6361 gggtagccaa cgctatgtcc tgatagcggt ccgccacacc cagccggcca cagtcgatga
     6421 atccagaaaa gcggccattt tccaccatga tattcggcaa gcaggcatcg ccatgggtca
     6481 cgacgagatc ctcgccgtcg ggcatgctcg ccttgagcct ggcgaacagt tcggctggcg
     6541 cgagcccctg atgctcttcg tccagatcat cctgatcgac aagaccggct tccatccgag
     6601 tacgtgctct ctcgatgcga tgtttcgctt ggtggtcgaa tgggcaggta gccggatcaa
     6661 gcgtatgcag ccgccgcatt gcatcagcca tgatggatac tttctcggca ggagcaaggt
     6721 gagatgacag gagatcctgc cccggcactt cgcccaatag cagccagtcc cttcccgctt
     6781 cagtgacaac gtcgagtaca gctgcgcaag gaacgcccgt cgtggccagc cacgatagcc
     6841 gcgctgcctc gtcttgcagt tcattcaggg caccggacag gtcggtcttg acaaaaagaa
     6901 ccgggcgccc ctgcgctgac agccggaaca cggcggcatc agagcagccg attgtctgtt
     6961 gtgcccagtc atagccgaat agcctctcca cccaagcggc cggagaacct gcgtgcaatc
     7021 catcttgttc aatcataata ttattgaagc atttatcagg gttcgtctcg tcccggtctc
     7081 ctcccatgca tg
//
