LOCUS       sequence-151767-        1071 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Cas9 protein with GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1071)
  AUTHORS   Pulido-Quetglas C, Aparicio-Prat E, Arnan C, Polidori T, Hermoso T, 
            Palumbo E, Ponomarenko J, Guigo R, Johnson R
  TITLE     Scalable Design of Paired CRISPR Guide RNAs for Genomic Deletion.
  JOURNAL   PLoS Comput Biol. 2017 Mar 2;13(3):e1005341. doi: 
            10.1371/journal.pcbi.1005341. eCollection 2017 Mar.
  PUBMED    28253259
REFERENCE   2  (bases 1 to 1071)
  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"; journalName: "PLoS Comput
            Biol."; date: "2017-03-2"; volume: "13(3):e1005341. doi"; pages: " 
            10.1371/journal.pcbi.1005341. eCollection 2017 Mar"
COMMENT     Sequence Label: pLentiCas9-T2A-GFP
FEATURES             Location/Qualifiers
     source          1..1071
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             53..100
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             101..124
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             134..187
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     regulatory      194..203
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(200..202,203..205,206..916)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKLSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(245..266)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(506..525)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     853..874
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             917..934
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             944..1000
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
ORIGIN
        1 catcaccggc ctgtacgaga cacggatcga cctgtctcag ctgggaggcg acaagcgacc
       61 tgccgccaca aagaaggctg gacaggctaa gaagaagaaa gattacaaag acgatgacga
      121 taagggatcc ggagagggca gaggaagtct gctaacatgc ggtgacgtcg aggagaatcc
      181 tggcccaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
      241 catcctggtc gagctggacg gcgacgtaaa cggccacaag ctcagcgtgt ccggcgaggg
      301 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
      361 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
      421 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
      481 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
      541 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
      601 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
      661 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
      721 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
      781 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
      841 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
      901 ggacgagctg tacaagcatc atcaccatca ccacggatcc ggcgcaacaa acttctctct
      961 gctgaaacaa gccggagatg tcgaagagaa tcctggaccg atggccaagc ctttgtctca
     1021 agaagaatcc accctcattg aaagagcaac ggctacaatc aacagcatcc c
//
