LOCUS       sequence-214588-        7484 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector expressing cell surface HA-tag and monomeric GFP 
            N-terminaly fused with auxin inducible degron.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7484)
  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 7484)
  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-CD5HA2-P2A-smAID-mClover
FEATURES             Location/Qualifiers
     source          1..7484
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             350..607
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    654..779
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1272..1505
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1690..1734
                     /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    2029..2146
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        2206..2509
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2510..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             2906..2932
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3020..3076
                     /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             join(3206..3208,3209..3922)
                     /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="VSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(3248..3269)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     3856..3877
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             3999..4632
                     /label=5' LTR
                     /note="5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(4681..4697)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4681..4697)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4694..4716)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4705..4721
                     /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(4729..4735,4736..4753,4754..4759))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4774..4795
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4912..4929)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      5072..5207
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     rep_origin      complement(5270..5858)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5350..5369)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6029..6820,6821..6889))
                     /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     6652..6671
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6890..6994)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7062..7080
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7118..7140)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7240..7259
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7453..7475
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7467..7484
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7468..7484
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gaattgatat ccttgatctg tggatctacc acacacaagg ctacttccct gattagcaga
       61 actacacacc agggccaggg gtcagatatc cactgacctt tggatggtgc tacaagctag
      121 taccagttga gccagataag gtagaagagg ccaataaagg agagaacacc agcttgttac
      181 accctgtgag cctgcatggg atggatgacc cggagagaga agtgttagag tggaggtttg
      241 acagccgcct agcatttcat cacgtggccc gagagctgca tccggagtac ttcaagaact
      301 gctgatatcg agcttgctac aagggacttt ccgctgggga ctttccaggg aggcgtggcc
      361 tgggcgggac tggggagtgg cgagccctca gatcctgcat ataagcagct gctttttgcc
      421 tgtactgggt ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg
      481 aacccactgc ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt
      541 ctgttgtgtg actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc
      601 tctagcagtg gcgcccgaac agggacttga aagcgaaagg gaaaccagag gagctctctc
      661 gacgcaggac tcggcttgct gaagcgcgca cggcaagagg cgaggggcgg cgactggtga
      721 gtacgccaaa aattttgact agcggaggct agaaggagag agatgggtgc gagagcgtca
      781 gtattaagcg ggggagaatt agatcgcgat gggaaaaaat tcggttaagg ccagggggaa
      841 agaaaaaata taaattaaaa catatagtat gggcaagcag ggagctagaa cgattcgcag
      901 ttaatcctgg cctgttagaa acatcagaag gctgtagaca aatactggga cagctacaac
      961 catcccttca gacaggatca gaagaactta gatcattata taatacagta gcaaccctct
     1021 attgtgtgca tcaaaggata gagataaaag acaccaagga agctttagac aagatagagg
     1081 aagagcaaaa caaaagtaag accaccgcac agcaagcggc cgctgatctt cagacctgga
     1141 ggaggagata tgagggacaa ttggagaagt gaattatata aatataaagt agtaaaaatt
     1201 gaaccattag gagtagcacc caccaaggca aagagaagag tggtgcagag agaaaaaaga
     1261 gcagtgggaa taggagcttt gttccttggg ttcttgggag cagcaggaag cactatgggc
     1321 gcagcgtcaa tgacgctgac ggtacaggcc agacaattat tgtctggtat agtgcagcag
     1381 cagaacaatt tgctgagggc tattgaggcg caacagcatc tgttgcaact cacagtctgg
     1441 ggcatcaagc agctccaggc aagaatcctg gctgtggaaa gatacctaaa ggatcaacag
     1501 ctcctgggga tttggggttg ctctggaaaa ctcatttgca ccactgctgt gccttggaat
     1561 gctagttgga gtaataaatc tctggaacag atttggaatc acacgacctg gatggagtgg
     1621 gacagagaaa ttaacaatta cacaagctta atacactcct taattgaaga atcgcaaaac
     1681 cagcaagaaa agaatgaaca agaattattg gaattagata aatgggcaag tttgtggaat
     1741 tggtttaaca taacaaattg gctgtggtat ataaaattat tcataatgat agtaggaggc
     1801 ttggtaggtt taagaatagt ttttgctgta ctttctatag tgaatagagt taggcaggga
     1861 tattcaccat tatcgtttca gacccacctc ccaaccccga ggggacccga caggcccgaa
     1921 ggaatagaag aagaaggtgg agagagagac agagacagat ccattcgatt agtgaacgga
     1981 tctcgacggt atcggaccct cgagacaaat ggcagtattc atccacaatt ttaaaagaaa
     2041 aggggggatt ggggggtaca gtgcagggga aagaatagta gacataatag caacagacat
     2101 acaaactaaa gaattacaaa aacaaattac aaaaattcaa aattttcggg tttattacag
     2161 ggacagcaga aatccacttt gggtccgata agcttgggag ttccgcgtta cataacttac
     2221 ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac
     2281 gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt
     2341 acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat
     2401 tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga
     2461 ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt
     2521 ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca
     2581 ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg
     2641 tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta
     2701 tataagcaga gctcgtttag tgaaccgtca gatcgcctgg agacgccatc cacgctgttt
     2761 tgacctccat agaagacacc gggaccgatc cagcctccga attcgcaggt cgaccaccat
     2821 gaagcgcttc ctcttcctcc tactcaccat cagcctcctg gttatggtac agatacaaac
     2881 tggactctca ggacaaaacg acacctaccc ttacgacgtg cccgactacg cctcagcatc
     2941 cagcaacata agcggaggca ttttcctttt cttcgtggcc aatgccataa tccacctctt
     3001 ctgcttcagt ggatccggcg caacaaactt ctctctgctg aaacaagccg gagatgtcga
     3061 agagaatcct ggaccggcca aacctccggc caaggcacaa gttgtgggat ggccaccggt
     3121 gagatcatac cggaagaacg tgatggtttc ctgccaaaaa tcaagcggtg gcccggaggc
     3181 ggcggcgttc gtgaaagtag ctagcgtgag caagggcgag gagctgttca ccggggtggt
     3241 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tccgcggcga
     3301 gggcgagggc gatgccacca acggcaagct gaccctgaag ttcatctgca ccaccggcaa
     3361 gctgcccgtg ccctggccca ccctcgtgac caccttcggc tacggcgtgg cctgcttcag
     3421 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
     3481 cgtccaggag cgcaccatct ctttcaagga cgacggtacc tacaagaccc gcgccgaggt
     3541 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     3601 ggacggcaac atcctggggc acaagctgga gtacaacttc aacagccaca acgtctatat
     3661 cacggccgac aagcagaaga acggcatcaa ggctaacttc aagatccgcc acaacgttga
     3721 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
     3781 cgtgctgctg cccgacaacc actacctgag ccatcagtcc aagctgagca aagaccccaa
     3841 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga ttacacatgg
     3901 catggacgag ctgtacaagt aagctcgagt acctttaaga ccaatgactt acaaggcagc
     3961 tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa ttcactccca
     4021 acgaagacaa gatatccttg atctgtggat ctaccacaca caaggctact tccctgattg
     4081 gcagaactac acaccagggc cagggatcag atatccactg acctttggat ggtgctacaa
     4141 gctagtacca gttgagcaag agaaggtaga agaagccaat gaaggagaga acacccgctt
     4201 gttacaccct gtgagcctgc atgggatgga tgacccggag agagaagtat tagagtggag
     4261 gtttgacagc cgcctagcat ttcatcacat ggcccgagag ctgcatccgg agtacttcaa
     4321 gaactgctga catcgagctt gctacaaggg actttccgct ggggactttc cagggaggcg
     4381 tggcctgggc gggactgggg agtggcgagc cctcagatgc tgcatataag cagctgcttt
     4441 ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac
     4501 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg
     4561 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga
     4621 aaatctctag caggatcctc tagagtcgac ctgcaggcat gcaagcttgg cgtaatcatg
     4681 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     4741 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     4801 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4861 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     4921 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     4981 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgcatc tcaattagtc
     5041 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     5101 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     5161 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag atgtgagcaa
     5221 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     5281 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     5341 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5401 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5461 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5521 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5581 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5641 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5701 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5761 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5821 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     5881 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     5941 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     6001 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     6061 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     6121 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     6181 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     6241 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     6301 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6361 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6421 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6481 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6541 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6601 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6661 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6721 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6781 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6841 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     6901 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     6961 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     7021 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     7081 cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     7141 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
     7201 cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag cagattgtac
     7261 tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca
     7321 tcaggcgcca ttcgccattc aggctgcgca actgttggga agggcgatcg gtgcgggcct
     7381 cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa
     7441 cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagt
//
