LOCUS       Exported                5177 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  BFPKRAB_PUFa in pmax-DEST expression vector.
ACCESSION   .
VERSION     .
KEYWORDS    pAC1451_pmax-BFPKRAB_PUFa
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5177)
  AUTHORS   Cheng AW, Jillette N, Lee P, Plaskon D, Fujiwara Y, Wang W, 
            Taghbalout A, Wang H
  TITLE     Casilio: a versatile CRISPR-Cas9-Pumilio hybrid for gene regulation 
            and genomic labeling.
  JOURNAL   Cell Res. 2016 Feb;26(2):254-7. doi: 10.1038/cr.2016.3. Epub 2016 
            Jan 15.
  PUBMED    26768771
REFERENCE   2  (bases 1 to 5177)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5177
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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"
     regulatory      1002..1011
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1014..1040
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1059..1079
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1086..1106
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1146..1841
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="SELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKVV
                     EGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTAT
                     QDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALKL
                     VGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYCD
                     LPSKLGHKLN"
     CDS             1887..2072
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEP"
     CDS             2115..2135
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     protein_bind    complement(3207..3231)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3284..3325
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3351..3370)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    3380..3461
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     3405..3424
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3659..4246)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3739..3758)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4327..5121)
                     /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(4438..4457)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5048..5067
                     /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 gcctatccct
     1021 atgacgtgcc cgattatgcc agcctgggca gcggctcccc caagaaaaaa cgcaaggtgg
     1081 aagatcctaa gaaaaagcgg aaagtggacg gcattggtag tgggagcaac ggcagcagcg
     1141 gatccagcga gctgattaag gagaacatgc acatgaagct gtacatggag ggcaccgtgg
     1201 acaaccatca cttcaagtgc acatccgagg gcgaaggcaa gccctacgag ggcacccaga
     1261 ccatgagaat caaggtggtc gagggcggcc ctctcccctt cgccttcgac atcctggcta
     1321 ctagcttcct ctacggcagc aagaccttca tcaaccacac ccagggcatc cccgacttct
     1381 tcaagcagtc cttccctgag ggcttcacat gggagagagt caccacatac gaagacgggg
     1441 gcgtgctgac cgctacccag gacaccagcc tccaggacgg ctgcctcatc tacaacgtca
     1501 agatcagagg ggtgaacttc acatccaacg gccctgtgat gcagaagaaa acactcggct
     1561 gggaggcctt caccgagacg ctgtaccccg ctgacggcgg cctggaaggc agaaacgaca
     1621 tggccctgaa gctcgtgggc gggagccatc tgatcgcaaa catcaagacc acatatagat
     1681 ccaagaaacc cgctaagaac ctcaagatgc ctggcgtcta ctatgtggac tacagactgg
     1741 aaagaatcaa ggaggccaac aacgagacct acgtcgagca gcacgaggtg gcagtggcca
     1801 gatactgcga cctccctagc aaactggggc acaagcttaa tggcggtggc ggagggatgg
     1861 atgctaagtc actaactgcc tggtcccgga cactggtgac cttcaaggat gtatttgtgg
     1921 acttcaccag ggaggagtgg aagctgctgg acactgctca gcagatcgtg tacagaaatg
     1981 tgatgctgga gaactataag aacctggttt ccttgggtta tcagcttact aagccagatg
     2041 tgatcctccg gttggagaag ggagaagagc ccggcggttc cggcggaggg tcggggccgg
     2101 ccgggatcct cccccccaag aaaaagagga aggtatctag aggccgcagc cgccttttgg
     2161 aagattttcg aaacaaccgg taccccaatt tacaactgcg ggagattgct ggacatataa
     2221 tggaattttc ccaagaccag catgggtcca gattcattca gctgaaactg gagcgtgcca
     2281 caccagctga gcgccagctt gtcttcaatg aaatcctcca ggctgcctac caactcatgg
     2341 tggatgtgtt tggtaattac gtcattcaga agttctttga atttggcagt cttgaacaga
     2401 agctggcttt ggcagaacgg attcgaggcc acgtcctgtc attggcacta cagatgtatg
     2461 gcagccgtgt tatcgagaaa gctcttgagt ttattccttc agaccagcag aatgagatgg
     2521 ttcgggaact agatggccat gtcttgaagt gtgtgaaaga tcagaatggc aatcacgtgg
     2581 ttcagaaatg cattgaatgt gtacagcccc agtctttgca atttatcatc gatgcgttta
     2641 agggacaggt atttgcctta tccacacatc cttatggctg ccgagtgatt cagagaatcc
     2701 tggagcactg tctccctgac cagacactcc ctattttaga ggagcttcac cagcacacag
     2761 agcagctggt acaggatcaa tatggaaatt atgtaatcca acatgtactg gagcacggtc
     2821 gtcctgagga taaaagcaaa attgtagcag aaatccgagg caatgtactt gtattgagtc
     2881 agcacaaatt tgcaagcaat gttgtggaga agtgtgttac tcacgcctca cgtacggagc
     2941 gcgctgtgct catcgatgag gtgtgcacca tgaacgacgg tccccacagt gccttataca
     3001 ccatgatgaa ggaccagtat gccaactacg tggtccagaa gatgattgac gtggcggagc
     3061 caggccagcg gaagatcgtc atgcataaga tccggcccca catcgcaact cttcgtaagt
     3121 acacctatgg caagcacatt ctggccaagc tggagaagta ctacatgaag aacggtgttg
     3181 acttagggtg attaattaag aattcgaccc agctttcttg tacaaagtgg ttgatatcca
     3241 gcacagtggc ggccgctcga gtctagaggg cccgcggttc gaaggtaagc ctatccctaa
     3301 ccctctcctc ggtctcgatt ctacgcgtac cggttagcaa ttgttttttc gatgagtttg
     3361 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     3421 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     3481 attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct
     3541 acaaatgtgg tacttaagag ggggagacca aagggcgaga cgttaaggcc tcacgtgaca
     3601 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     3661 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     3721 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     3781 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     3841 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     3901 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     3961 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     4021 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     4081 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
     4141 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac cacgctggta gcggtggttt
     4201 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4261 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4321 gccgtctcag aagaactcgt caagaaggcg atagaaggcg atgcgctgcg aatcgggagc
     4381 ggcgataccg taaagcacga ggaagcggtc agcccattcg ccgccaagct cttcagcaat
     4441 atcacgggta gccaacgcta tgtcctgata gcggtccgcc acacccagcc ggccacagtc
     4501 gatgaatcca gaaaagcggc cattttccac catgatattc ggcaagcagg catcgccatg
     4561 ggtcacgacg agatcctcgc cgtcgggcat gctcgccttg agcctggcga acagttcggc
     4621 tggcgcgagc ccctgatgct cttcgtccag atcatcctga tcgacaagac cggcttccat
     4681 ccgagtacgt gctcgctcga tgcgatgttt cgcttggtgg tcgaatgggc aggtagccgg
     4741 atcaagcgta tgcagccgcc gcattgcatc agccatgatg gatactttct cggcaggagc
     4801 aaggtgagat gacaggagat cctgccccgg cacttcgccc aatagcagcc agtcccttcc
     4861 cgcttcagtg acaacgtcga gcacagctgc gcaaggaacg cccgtcgtgg ccagccacga
     4921 tagccgcgct gcctcgtctt gcagttcatt cagggcaccg gacaggtcgg tcttgacaaa
     4981 aagaaccggg cgcccctgcg ctgacagccg gaacacggcg gcatcagagc agccgattgt
     5041 ctgttgtgcc cagtcatagc cgaatagcct ctccacccaa gcggccggag aacctgcgtg
     5101 caatccatct tgttcaatca taatattatt gaagcattta tcagggttcg tctcgtcccg
     5161 gtctcctccc atgcatg
//
