LOCUS       sequence-232813-        8810 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For LIC cloning with C-terminal V5 tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8810)
  AUTHORS   Cheng LC, Baboo S, Lindsay C, Brusman L, Martinez-Bartolome S, Tapia
            O, Zhang X, Yates JR 3rd, Gerace L
  TITLE     Identification of new transmembrane proteins concentrated at the 
            nuclear envelope using organellar proteomics of mesenchymal cells.
  JOURNAL   Nucleus. 2019 Dec;10(1):126-143. doi: 10.1080/19491034.2019.1618175.
  PUBMED    31142202
REFERENCE   2  (bases 1 to 8810)
  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"; doi: 
            "10.1080/19491034.2019"; journalName: "Nucleus"; date: "2019-12"; 
            volume: "10"; issue: "1"; pages: "126-143"
COMMENT     Sequence Label: pLV-EF1a-LIC-V5
FEATURES             Location/Qualifiers
     source          1..8810
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        78..289
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     intron          290..1228
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1185..1205
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             1283..1324
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     misc_feature    1461..2010
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1605..1622)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1832..1851
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2013..2612
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(2013..2032)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2509..2529
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    2636..3224
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2689..2709)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3107..3118)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             3431..4064
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(4192..4208)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4192..4208)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4205..4227)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4216..4232
                     /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(4240..4246,4247..4264,4265..4270))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4285..4306
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4423..4440)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4594..5182)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4674..4693)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5353..6144,6145..6213))
                     /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     5976..5995
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6214..6318)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    6366..6500
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6403..6422)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6457..6476
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     LTR             6668..7301
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    7348..7473
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7970..8203
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             8388..8432
                     /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    8695..8810
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
ORIGIN
        1 tcgggtttat tacagggaca gcagagatcc agtttatcga tgaggccctt tcgtcttcac
       61 tcgaggtgcc cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg
      121 aggggtcggc aattgaaccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga
      181 tgtcgtgtac tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt
      241 agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg taagtgccgt
      301 gtgtggttcc cgcgggcctg gcctctttac gggttatggc ccttgcgtgc cttgaattac
      361 ttccacctgg ctgcagtacg tgattcttga tcccgagctt cgggttggaa gtgggtggga
      421 gagttcgagg ccttgcgctt aaggagcccc ttcgcctcgt gcttgagttg aggcctggcc
      481 tgggcgctgg ggccgccgcg tgcgaatctg gtggcacctt cgcgcctgtc tcgctgcttt
      541 cgataagtct ctagccattt aaaatttttg atgacctgct gcgacgcttt ttttctggca
      601 agatagtctt gtaaatgcgg gccaagatct gcacactggt atttcggttt ttggggccgc
      661 gggcggcgac ggggcccgtg cgtcccagcg cacatgttcg gcgaggcggg gcctgcgagc
      721 gcggccaccg agaatcggac gggggtagtc tcaagctggc cggcctgctc tggtgcctgg
      781 cctcgcgccg ccgtgtatcg ccccgccctg ggcggcaagg ctggcccggt cggcaccagt
      841 tgcgtgagcg gaaagatggc cgcttcccgg ccctgctgca gggagctcaa aatggaggac
      901 gcggcgctcg ggagagcggg cgggtgagtc acccacacaa aggaaaaggg cctttccgtc
      961 ctcagccgtc gcttcatgtg actccacgga gtaccgggcg ccgtccaggc acctcgatta
     1021 gttctcgagc ttttggagta cgtcgtcttt aggttggggg gaggggtttt atgcgatgga
     1081 gtttccccac actgagtggg tggagactga agttaggcca gcttggcact tgatgtaatt
     1141 ctccttggaa tttgcccttt ttgagtttgg atcttggttc attctcaagc ctcagacagt
     1201 ggttcaaagt ttttttcttc catttcaggt gtcgtgagga tccgccatcg gcccgcccgg
     1261 gccgggcggg ggtggcgggg gaggtaagcc tatccctaac cctctcctcg gtctcgattc
     1321 tacgtgagtt taaacttcga aacgcgtgaa ttcctcgagg gcggccgctc tagagtcgac
     1381 gggccgcggt aacaattgtt aactaactta agctagcaac ggtttccctc tagcgggatc
     1441 aattccgccc ccccccccta acgttactgg ccgaagccgc ttggaataag gccggtgtgc
     1501 gtttgtctat atgttatttt ccaccatatt gccgtctttt ggcaatgtga gggcccggaa
     1561 acctggccct gtcttcttga cgagcattcc taggggtctt tcccctctcg ccaaaggaat
     1621 gcaaggtctg ttgaatgtcg tgaaggaagc agttcctctg gaagcttctt gaagacaaac
     1681 aacgtctgta gcgacccttt gcaggcagcg gaacccccca cctggcgaca ggtgcctctg
     1741 cggccaaaag ccacgtgtat aagatacacc tgcaaaggcg gcacaacccc agtgccacgt
     1801 tgtgagttgg atagttgtgg aaagagtcaa atggctctcc tcaagcgtat tcaacaaggg
     1861 gctgaaggat gcccagaagg taccccattg tatgggatct gatctggggc ctcggtgcac
     1921 atgctttaca tgtgtttagt cgaggttaaa aaaacgtcta ggccccccga accacgggga
     1981 cgtggttttc ctttgaaaaa cacgataata ccatgaccga gtacaagccc acggtgcgcc
     2041 tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg ttcgccgact
     2101 accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc accgagctgc
     2161 aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg
     2221 gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg gtgttcgccg
     2281 agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag caacagatgg
     2341 aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc accgtcggcg
     2401 tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg
     2461 cggccgagcg cgccggggtg cccgccttcc tggagacctc cgcgccccgc aacctcccct
     2521 tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca
     2581 cctggtgcat gacccgcaag cccggtgcct gacccgggcg gggcgcgtct ggaacaatca
     2641 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     2701 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     2761 tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc
     2821 cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg
     2881 gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc
     2941 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     3001 cactgacaat tccgtggtgt tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg
     3061 tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc
     3121 agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct
     3181 tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcctggaa ttaattctgc
     3241 agtcgagacc tagaaaaaca tggagcaatc acaagtagca atacagcagc taccaatgct
     3301 gattgtgcct ggctagaagc acaagaggag gaggaggtgg gttttccagt cacacctcag
     3361 gtacctttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa
     3421 aagaggggac tggaagggct aattcactcc caacgaagac aagatatcct tgatctgtgg
     3481 atctaccaca cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc
     3541 agatatccac tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta
     3601 gaagaggcca ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg
     3661 gatgacccgg agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac
     3721 gtggcccgag agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag
     3781 ggactttccg ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga
     3841 gccctcagat cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga
     3901 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata
     3961 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta
     4021 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtagta gttcatgtca
     4081 tcttattatt cagtatttat aacttgcaaa gaaatgaata tcagagagtg agaggccttg
     4141 acattgctag cgtttaccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     4201 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     4261 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     4321 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     4381 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     4441 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     4501 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     4561 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4621 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4681 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4741 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     4801 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     4861 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     4921 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     4981 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     5041 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     5101 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     5161 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5221 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5281 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     5341 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     5401 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     5461 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     5521 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     5581 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5641 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5701 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     5761 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     5821 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     5881 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     5941 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6001 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6061 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6121 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6181 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     6241 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6301 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg
     6361 agatcaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     6421 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     6481 tatcttatca tgtctggatc aactggataa ctcaagctaa ccaaaatcat cccaaacttc
     6541 ccaccccata ccctattacc actgccaatt acctgtggtt tcatttactc taaacctgtg
     6601 attcctctga attattttca ttttaaagaa attgtatttg ttaaatatgt actacaaact
     6661 tagtagttgg aagggctaat tcactcccaa agaagacaag atatccttga tctgtggatc
     6721 taccacacac aaggctactt ccctgattag cagaactaca caccagggcc aggggtcaga
     6781 tatccactga cctttggatg gtgctacaag ctagtaccag ttgagccaga taaggtagaa
     6841 gaggccaata aaggagagaa caccagcttg ttacaccctg tgagcctgca tgggatggat
     6901 gacccggaga gagaagtgtt agagtggagg tttgacagcc gcctagcatt tcatcacgtg
     6961 gcccgagagc tgcatccgga gtacttcaag aactgctgat atcgagcttg ctacaaggga
     7021 ctttccgctg gggactttcc agggaggcgt ggcctgggcg ggactgggga gtggcgagcc
     7081 ctcagatcct gcatataagc agctgctttt tgcctgtact gggtctctct ggttagacca
     7141 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag
     7201 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag
     7261 atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc gaacagggac
     7321 ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct tgctgaagcg
     7381 cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt gactagcgga
     7441 ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag aattagatcg
     7501 cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt aaaacatata
     7561 gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt agaaacatca
     7621 gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg atcagaagaa
     7681 cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag gatagagata
     7741 aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag taagaccacc
     7801 gcacagcaag cggccggccg ctgatcttca gacctggagg aggagatatg agggacaatt
     7861 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
     7921 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
     7981 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg
     8041 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
     8101 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
     8161 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
     8221 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
     8281 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca
     8341 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag
     8401 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc
     8461 tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt
     8521 ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga
     8581 cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag
     8641 agagagacag agacagatcc attcgattag tgaacggatc tcgacggtat cgcctttaaa
     8701 agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca
     8761 gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
//
