LOCUS       sequence-208136-        7555 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector expressing cell surface Flag-tag and monomeric GFP
            C-terminaly fused with auxin inducible degron.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7555)
  AUTHORS   Zotova A, Pichugin A, Atemasova A, Knyazhanskaya E, Lopatukhina E, 
            Mitkin N, Holmuhamedov E, Gottikh M, Kuprash D, Filatov A, Mazurov D
  TITLE     Isolation of gene-edited cells via knock-in of short 
            glycophosphatidylinositol-anchored epitope tags.
  JOURNAL   Sci Rep. 2019 Feb 28;9(1):3132. doi: 10.1038/s41598-019-40219-z.
  PUBMED    30816313
REFERENCE   2  (bases 1 to 7555)
  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: "Sci Rep."; 
            date: "2019-02-28"; volume: "9(1):3132. doi"; pages: " 
            10.1038/s41598-019-40219-z"
COMMENT     Sequence Label: pUCHR-mClover-smAID-P2A-CD5Flag2-bglpA
FEATURES             Location/Qualifiers
     source          1..7555
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             352..609
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    656..781
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1274..1507
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1692..1736
                     /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    2031..2148
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        2208..2511
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2512..2713
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2664..2684
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2710..2734
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(2827..2829,2830..2832,2833..3543)
                     /codon_start=1
                     /product="bright green-yellow fluorescent protein derived 
                     from mClover2, with improved photostability (Bajar et al., 
                     2003)"
                     /label=mClover3
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(2872..2893)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     3480..3501
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3679..3735
                     /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"
     CDS             3820..3843
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    3924..3972
                     /note="synthetic polyadenylation signal"
     LTR             4069..4702
                     /label=5' LTR
                     /note="5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(4751..4767)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4751..4767)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4764..4786)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4775..4791
                     /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)."
     promoter        complement(join(4799..4805,4806..4823,4824..4829))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4844..4865
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4982..4999)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      5142..5277
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     rep_origin      complement(5341..5929)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5421..5440)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6100..6891,6892..6960))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6723..6742
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6961..7065)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7133..7151
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7189..7211)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7311..7330
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7524..7546
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7538..7555
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7539..7555
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gaattaagat atccttgatc tgtggatcta ccacacacaa ggctacttcc ctgattagca
       61 gaactacaca ccagggccag gggtcagata tccactgacc tttggatggt gctacaagct
      121 agtaccagtt gagccagata aggtagaaga ggccaataaa ggagagaaca ccagcttgtt
      181 acaccctgtg agcctgcatg ggatggatga cccggagaga gaagtgttag agtggaggtt
      241 tgacagccgc ctagcatttc atcacgtggc ccgagagctg catccggagt acttcaagaa
      301 ctgctgatat cgagcttgct acaagggact ttccgctggg gactttccag ggaggcgtgg
      361 cctgggcggg actggggagt ggcgagccct cagatcctgc atataagcag ctgctttttg
      421 cctgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
      481 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
      541 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
      601 tctctagcag tggcgcccga acagggactt gaaagcgaaa gggaaaccag aggagctctc
      661 tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt
      721 gagtacgcca aaaattttga ctagcggagg ctagaaggag agagatgggt gcgagagcgt
      781 cagtattaag cgggggagaa ttagatcgcg atgggaaaaa attcggttaa ggccaggggg
      841 aaagaaaaaa tataaattaa aacatatagt atgggcaagc agggagctag aacgattcgc
      901 agttaatcct ggcctgttag aaacatcaga aggctgtaga caaatactgg gacagctaca
      961 accatccctt cagacaggat cagaagaact tagatcatta tataatacag tagcaaccct
     1021 ctattgtgtg catcaaagga tagagataaa agacaccaag gaagctttag acaagataga
     1081 ggaagagcaa aacaaaagta agaccaccgc acagcaagcg gccgctgatc ttcagacctg
     1141 gaggaggaga tatgagggac aattggagaa gtgaattata taaatataaa gtagtaaaaa
     1201 ttgaaccatt aggagtagca cccaccaagg caaagagaag agtggtgcag agagaaaaaa
     1261 gagcagtggg aataggagct ttgttccttg ggttcttggg agcagcagga agcactatgg
     1321 gcgcagcgtc aatgacgctg acggtacagg ccagacaatt attgtctggt atagtgcagc
     1381 agcagaacaa tttgctgagg gctattgagg cgcaacagca tctgttgcaa ctcacagtct
     1441 ggggcatcaa gcagctccag gcaagaatcc tggctgtgga aagataccta aaggatcaac
     1501 agctcctggg gatttggggt tgctctggaa aactcatttg caccactgct gtgccttgga
     1561 atgctagttg gagtaataaa tctctggaac agatttggaa tcacacgacc tggatggagt
     1621 gggacagaga aattaacaat tacacaagct taatacactc cttaattgaa gaatcgcaaa
     1681 accagcaaga aaagaatgaa caagaattat tggaattaga taaatgggca agtttgtgga
     1741 attggtttaa cataacaaat tggctgtggt atataaaatt attcataatg atagtaggag
     1801 gcttggtagg tttaagaata gtttttgctg tactttctat agtgaataga gttaggcagg
     1861 gatattcacc attatcgttt cagacccacc tcccaacccc gaggggaccc gacaggcccg
     1921 aaggaataga agaagaaggt ggagagagag acagagacag atccattcga ttagtgaacg
     1981 gatctcgacg gtatcggacc ctcgagacaa atggcagtat tcatccacaa ttttaaaaga
     2041 aaagggggga ttggggggta cagtgcaggg gaaagaatag tagacataat agcaacagac
     2101 atacaaacta aagaattaca aaaacaaatt acaaaaattc aaaattttcg ggtttattac
     2161 agggacagca gaaatccact ttgggtccga taagcttggg agttccgcgt tacataactt
     2221 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
     2281 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
     2341 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
     2401 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttacgg
     2461 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
     2521 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacgggattt ccaagtctcc
     2581 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac ttccaaaatg
     2641 tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta
     2701 tataagcaga gctcgtttag tgaaccgtca gatcgcctgg agacgccatc cacgctgttt
     2761 tgacctccat agaagacacc gggaccgatc cagcctccca agcttcgaat tcgcaggtcg
     2821 accaccatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag
     2881 ctggacggcg acgtaaacgg ccacaagttc agcgtccgcg gcgagggcga gggcgatgcc
     2941 accaacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
     3001 cccaccctcg tgaccacctt cggctacggc gtggcctgct tcagccgcta ccccgaccac
     3061 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc
     3121 atctctttca aggacgacgg tacctacaag acccgcgccg aggtgaagtt cgagggcgac
     3181 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
     3241 gggcacaagc tggagtacaa cttcaacagc cacaacgtct atatcacggc cgacaagcag
     3301 aagaacggca tcaaggctaa cttcaagatc cgccacaacg ttgaggacgg cagcgtgcag
     3361 ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac
     3421 aaccactacc tgagccatca gtccaagctg agcaaagacc ccaacgagaa gcgcgatcac
     3481 atggtcctgc tggagttcgt gaccgccgcc gggattacac atggcatgga cgagctgtac
     3541 aagccagcca aacctccggc caaggcacaa gttgtgggat ggccaccggt gagatcatac
     3601 cggaagaacg tgatggtttc ctgccaaaaa tcaagcggtg gcccggaggc ggcggcgttc
     3661 gtgaaagtag gatccggcgc aacaaacttc tctctgctga aacaagccgg agatgtcgaa
     3721 gagaatcctg gaccgaagcg cttcctcttc ctcctactca ccatcagcct cctggttatg
     3781 gtacagatac aaactggact ctcaggacaa aacgacaccg actacaaaga cgatgacgac
     3841 aagtcagcat ccagcaacat aagcggaggc attttccttt tcttcgtggc caatgccata
     3901 atccacctct tctgcttcag ttgaataaaa tatctttatt ttcattacat ctgtgtgttg
     3961 gttttttgtg tggtaccgca tcttgctgaa aaactcgagt acctttaaga ccaatgactt
     4021 acaaggcagc tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa
     4081 ttcactccca acgaagacaa gatatccttg atctgtggat ctaccacaca caaggctact
     4141 tccctgattg gcagaactac acaccagggc cagggatcag atatccactg acctttggat
     4201 ggtgctacaa gctagtacca gttgagcaag agaaggtaga agaagccaat gaaggagaga
     4261 acacccgctt gttacaccct gtgagcctgc atgggatgga tgacccggag agagaagtat
     4321 tagagtggag gtttgacagc cgcctagcat ttcatcacat ggcccgagag ctgcatccgg
     4381 agtacttcaa gaactgctga catcgagctt gctacaaggg actttccgct ggggactttc
     4441 cagggaggcg tggcctgggc gggactgggg agtggcgagc cctcagatgc tgcatataag
     4501 cagctgcttt ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc
     4561 tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa
     4621 gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag
     4681 tcagtgtgga aaatctctag caggatcctc tagagtcgac ctgcaggcat gcaagcttgg
     4741 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     4801 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     4861 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     4921 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     4981 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     5041 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgcatc
     5101 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     5161 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     5221 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     5281 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     5341 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     5401 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     5461 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     5521 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     5581 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     5641 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     5701 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     5761 taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac
     5821 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     5881 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     5941 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     6001 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     6061 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     6121 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     6181 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     6241 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     6301 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     6361 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     6421 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     6481 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     6541 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     6601 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     6661 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     6721 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     6781 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     6841 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     6901 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     6961 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     7021 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     7081 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     7141 tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     7201 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     7261 tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga
     7321 gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag
     7381 aaaataccgc atcaggcgcc attcgccatt caggctgcgc aactgttggg aagggcgatc
     7441 ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt
     7501 aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagt
//
