LOCUS       sequence-193737-        7805 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  to epress PCP-GFP fusion protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7805)
  AUTHORS   Qin P, Parlak M, Kuscu C, Bandaria J, Mir M, Szlachta K, Singh R, 
            Darzacq X, Yildiz A, Adli M
  TITLE     Live cell imaging of low- and non-repetitive chromosome loci using 
            CRISPR-Cas9.
  JOURNAL   Nat Commun. 2017 Mar 14;8:14725. doi: 10.1038/ncomms14725.
  PUBMED    28290446
REFERENCE   2  (bases 1 to 7805)
  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: "Nat 
            Commun."; date: "2017-03-14"; volume: "8:14725. doi"; pages: " 
            10.1038/ncomms14725"
COMMENT     Sequence Label: PCP-GFP
FEATURES             Location/Qualifiers
     source          1..7805
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..399
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     regulatory      402..411
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             414..434
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             438..464
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             438..464
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             465..476
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     regulatory      1239..1248
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1245..1247,1248..1250,1251..1964)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1245..1247,1248..1250,1251..1961)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1290..1311)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1551..1570)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1898..1919
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1978..2566
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2031..2051)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2449..2460)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             2641..2874
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(2993..3011)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3079..3183
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3184..3252,3253..4044)
                     /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(3402..3421)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4215..4803
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4704..4723
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4957..4974
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5214..5235
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5250..5255,5256..5273,5274..5280)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5288..5304
                     /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     5293..5315
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5312..5328
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5312..5328
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5378..5397
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     LTR             5589..6222
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    6269..6394
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6891..7124
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7309..7353
                     /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    7646..7763
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc cctcctcacg gcgagcgctg
       61 ccacgtcaga cgaagggcgc agcgagcgtc ctgatccttc cgcccggacg ctcaggacag
      121 cggcccgctg ctcataagac tcggccttag aaccccagta tcagcagaag gacattttag
      181 gacgggactt gggtgactct agggcactgg ttttctttcc agagagcgga acaggcgagg
      241 aaaagtagtc ccttctcggc gattctgcgg agggatctcc gtggggcggt gaacgccgat
      301 gattatataa ggacgcgccg ggtgtggcac agctagttcc gtcgcagccg ggatttgggt
      361 cgcggttctt gtttgtggat cgctgtgatc gtcacttggc ggccgccatg ggcccaaaaa
      421 agaaaagaaa agttggctac ccctacgacg tgcccgacta cgccatcgaa ggccgccata
      481 tgctagcctc caaaaccatc gttctttcgg tcggcgaggc tactcgcact ctgactgaga
      541 tccagtccac cgcagaccgt cagatcttcg aagagaaggt cgggcctctg gtgggtcggc
      601 tgcgcctcac ggcttcgctc cgtcaaaacg gagccaagac cgcgtatcgc gtcaacctaa
      661 aactggatca ggcggacgtc gttgattccg gacttccgaa agtgcgctac actcaggtat
      721 ggtcgcacga cgtgacaatc gttgcgaata gcaccgaggc ctcgcgcaaa tcgttgtacg
      781 atttgaccaa gtccctcgtc gcgacctcgc aggtcgaaga tcttgtcgtc aaccttgtgc
      841 cgctgggccg tgcggatccg ctagcctcca aaaccatcgt tctttcggtc ggcgaggcta
      901 ctcgcactct gactgagatc cagtccaccg cagaccgtca gatcttcgaa gagaaggtcg
      961 ggcctctggt gggtcggctg cgcctcacgg cttcgctccg tcaaaacgga gccaagaccg
     1021 cgtatcgcgt caacctaaaa ctggatcagg cggacgtcgt tgattccgga cttccgaaag
     1081 tgcgctacac tcaggtatgg tcgcacgacg tgacaatcgt tgcgaatagc accgaggcct
     1141 cgcgcaaatc gttgtacgat ttgaccaagt ccctcgtcgc gacctcgcag gtcgaagatc
     1201 ttgtcgtcaa ccttgtgccg ctgggccgtc caccggtcgc caccatggtg agcaagggcg
     1261 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     1321 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     1381 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga
     1441 cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca
     1501 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     1561 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     1621 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     1681 acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact
     1741 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     1801 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt
     1861 ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     1921 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaaatcgat agatcctaat
     1981 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct
     2041 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg
     2101 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg
     2161 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt
     2221 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt
     2281 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg
     2341 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc
     2401 tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat
     2461 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc
     2521 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcctga gatcctttaa
     2581 gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa aaggggggac
     2641 tggaagggct aattcactcc caacgaagac aagatctgct ttttgcttgt actgggtctc
     2701 tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta
     2761 agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact
     2821 ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtagta
     2881 gttcatgtca tcttattatt cagtatttat aacttgcaaa gaaatgaata tcagagagtg
     2941 agaggcccgg gttaattaag gaaagggcta gatcattctt gaagacgaaa gggcctcgtg
     3001 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc
     3061 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     3121 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     3181 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     3241 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     3301 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     3361 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     3421 tcccgtgttg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     3481 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     3541 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     3601 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     3661 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     3721 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     3781 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     3841 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     3901 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     3961 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     4021 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     4081 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     4141 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     4201 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     4261 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     4321 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     4381 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     4441 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     4501 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     4561 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     4621 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     4681 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     4741 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     4801 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     4861 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     4921 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     4981 gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     5041 aagctcatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg
     5101 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
     5161 ccgtggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg
     5221 agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg
     5281 tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacat gattacgaat
     5341 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     5401 gtatcttatc atgtctggat caactggata actcaagcta accaaaatca tcccaaactt
     5461 cccaccccat accctattac cactgccaat tacctgtggt ttcatttact ctaaacctgt
     5521 gattcctctg aattattttc attttaaaga aattgtattt gttaaatatg tactacaaac
     5581 ttagtagttg gaagggctaa ttcactccca aagaagacaa gatatccttg atctgtggat
     5641 ctaccacaca caaggctact tccctgatta gcagaactac acaccagggc caggggtcag
     5701 atatccactg acctttggat ggtgctacaa gctagtacca gttgagccag ataaggtaga
     5761 agaggccaat aaaggagaga acaccagctt gttacaccct gtgagcctgc atgggatgga
     5821 tgacccggag agagaagtgt tagagtggag gtttgacagc cgcctagcat ttcatcacgt
     5881 ggcccgagag ctgcatccgg agtacttcaa gaactgctga tatcgagctt gctacaaggg
     5941 actttccgct ggggactttc cagggaggcg tggcctgggc gggactgggg agtggcgagc
     6001 cctcagatcc tgcatataag cagctgcttt ttgcctgtac tgggtctctc tggttagacc
     6061 agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa
     6121 gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga
     6181 gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc cgaacaggga
     6241 cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc ttgctgaagc
     6301 gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt tgactagcgg
     6361 aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga gaattagatc
     6421 gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat taaaacatat
     6481 agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt tagaaacatc
     6541 agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag gatcagaaga
     6601 acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa ggatagagat
     6661 aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa gtaagaccac
     6721 cgcacagcaa gcggccggcc gctgatcttc agacctggag gaggagatat gagggacaat
     6781 tggagaagtg aattatataa atataaagta gtaaaaattg aaccattagg agtagcaccc
     6841 accaaggcaa agagaagagt ggtgcagaga gaaaaaagag cagtgggaat aggagctttg
     6901 ttccttgggt tcttgggagc agcaggaagc actatgggcg cagcgtcaat gacgctgacg
     6961 gtacaggcca gacaattatt gtctggtata gtgcagcagc agaacaattt gctgagggct
     7021 attgaggcgc aacagcatct gttgcaactc acagtctggg gcatcaagca gctccaggca
     7081 agaatcctgg ctgtggaaag atacctaaag gatcaacagc tcctggggat ttggggttgc
     7141 tctggaaaac tcatttgcac cactgctgtg ccttggaatg ctagttggag taataaatct
     7201 ctggaacaga tttggaatca cacgacctgg atggagtggg acagagaaat taacaattac
     7261 acaagcttaa tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa
     7321 gaattattgg aattagataa atgggcaagt ttgtggaatt ggtttaacat aacaaattgg
     7381 ctgtggtata taaaattatt cataatgata gtaggaggct tggtaggttt aagaatagtt
     7441 tttgctgtac tttctatagt gaatagagtt aggcagggat attcaccatt atcgtttcag
     7501 acccacctcc caaccccgag gggacccgac aggcccgaag gaatagaaga agaaggtgga
     7561 gagagagaca gagacagatc cattcgatta gtgaacggat ctcgacggta tcgccgaatt
     7621 cacaaatggc agtattcatc cacaatttta aaagaaaagg ggggattggg gggtacagtg
     7681 caggggaaag aatagtagac ataatagcaa cagacataca aactaaagaa ttacaaaaac
     7741 aaattacaaa aattcaaaat tttcgggttt attacaggga cagcagagat ccagtttgga
     7801 ctagt
//
