LOCUS       sequence-227951-        8858 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For LIC cloning with N-terminal V5 tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8858)
  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 8858)
  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-V5-LIC
FEATURES             Location/Qualifiers
     source          1..8858
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..634
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1721..1765
                     /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    2028..2143
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
     promoter        2221..2432
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     intron          2433..3371
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3328..3348
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      3387..3396
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3396..3437
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     misc_feature    3605..4154
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3749..3766)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3976..3995
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4157..4756
                     /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(4157..4176)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4653..4673
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    4780..5368
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4833..4853)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5251..5262)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             5576..6209
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(6338..6354)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6338..6354)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6351..6373)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6362..6378
                     /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(6386..6392,6393..6410,6411..6416))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6431..6452
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6569..6586)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6740..7325)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6820..6839)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7496..8287,8288..8356))
                     /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     8119..8138
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8357..8461)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    8509..8643
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8546..8565)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8600..8619
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccttt aaaagaaaag
     2041 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2101 aaactaaaga actacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
     2161 acagcagaga tccagtttat cgatgaggcc ctttcgtctt cactcgaggt gcccgtcagt
     2221 gggcagagcg cacatcgccc acagtccccg agaagttggg gggaggggtc ggcaattgaa
     2281 ccggtgccta gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg tactggctcc
     2341 gcctttttcc cgagggtggg ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc
     2401 tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc cgtgtgtggt tcccgcgggc
     2461 ctggcctctt tacgggttat ggcccttgcg tgccttgaat tacttccacc tggctgcagt
     2521 acgtgattct tgatcccgag cttcgggttg gaagtgggtg ggagagttcg aggccttgcg
     2581 cttaaggagc cccttcgcct cgtgcttgag ttgaggcctg gcctgggcgc tggggccgcc
     2641 gcgtgcgaat ctggtggcac cttcgcgcct gtctcgctgc tttcgataag tctctagcca
     2701 tttaaaattt ttgatgacct gctgcgacgc tttttttctg gcaagatagt cttgtaaatg
     2761 cgggccaaga tctgcacact ggtatttcgg tttttggggc cgcgggcggc gacggggccc
     2821 gtgcgtccca gcgcacatgt tcggcgaggc ggggcctgcg agcgcggcca ccgagaatcg
     2881 gacgggggta gtctcaagct ggccggcctg ctctggtgcc tggcctcgcg ccgccgtgta
     2941 tcgccccgcc ctgggcggca aggctggccc ggtcggcacc agttgcgtga gcggaaagat
     3001 ggccgcttcc cggccctgct gcagggagct caaaatggag gacgcggcgc tcgggagagc
     3061 gggcgggtga gtcacccaca caaaggaaaa gggcctttcc gtcctcagcc gtcgcttcat
     3121 gtgactccac ggagtaccgg gcgccgtcca ggcacctcga ttagttctcg agcttttgga
     3181 gtacgtcgtc tttaggttgg ggggaggggt tttatgcgat ggagtttccc cacactgagt
     3241 gggtggagac tgaagttagg ccagcttggc acttgatgta attctccttg gaatttgccc
     3301 tttttgagtt tggatcttgg ttcattctca agcctcagac agtggttcaa agtttttttc
     3361 ttccatttca ggtgtcgtga ggatccgccg ccatgggtaa gcctatccct aaccctctcc
     3421 tcggtctcga ttctacgggg ggcggagggg gtgggcccgg gccgcgacga tgtttaaact
     3481 tcgaaacgcg tgaattcctc gagggcggcc gctctagagt cgacgggccg cggtaacaat
     3541 tgttaactaa cttaagctag caacggtttc cctctagcgg gatcaattcc gccccccccc
     3601 cctaacgtta ctggccgaag ccgcttggaa taaggccggt gtgcgtttgt ctatatgtta
     3661 ttttccacca tattgccgtc ttttggcaat gtgagggccc ggaaacctgg ccctgtcttc
     3721 ttgacgagca ttcctagggg tctttcccct ctcgccaaag gaatgcaagg tctgttgaat
     3781 gtcgtgaagg aagcagttcc tctggaagct tcttgaagac aaacaacgtc tgtagcgacc
     3841 ctttgcaggc agcggaaccc cccacctggc gacaggtgcc tctgcggcca aaagccacgt
     3901 gtataagata cacctgcaaa ggcggcacaa ccccagtgcc acgttgtgag ttggatagtt
     3961 gtggaaagag tcaaatggct ctcctcaagc gtattcaaca aggggctgaa ggatgcccag
     4021 aaggtacccc attgtatggg atctgatctg gggcctcggt gcacatgctt tacatgtgtt
     4081 tagtcgaggt taaaaaaacg tctaggcccc ccgaaccacg gggacgtggt tttcctttga
     4141 aaaacacgat aataccatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga
     4201 cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca
     4261 caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac
     4321 gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt
     4381 ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat
     4441 ggccgagttg agcggttccc ggctggccgc gcagcaacag atggaaggcc tcctggcgcc
     4501 gcaccggccc aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca
     4561 gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg
     4621 ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc cccttctacg agcggctcgg
     4681 cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg
     4741 caagcccggt gcctgacccg ggcggggcgc gtctggaaca atcaacctct ggattacaaa
     4801 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac
     4861 gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc
     4921 ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt
     4981 ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc
     5041 tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc
     5101 gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg
     5161 gtgttgtcgg ggaagctgac gtcctttcca tggctgctcg cctgtgttgc cacctggatt
     5221 ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc
     5281 cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt
     5341 cggatctccc tttgggccgc ctccccgcct ggaattaatt ctgcagtcga gacctagaaa
     5401 aacatggagc aatcacaagt agcaatacag cagctaccaa tgctgattgt gcctggctag
     5461 aagcacaaga ggaggaggag gtgggttttt ccagtcacac ctcaggtacc tttaagacca
     5521 atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaagag gggactggaa
     5581 gggctaattc actcccaacg aagacaagat atccttgatc tgtggatcta ccacacacaa
     5641 ggctacttcc ctgattagca gaactacaca ccagggccag gggtcagata tccactgacc
     5701 tttggatggt gctacaagct agtaccagtt gagccagata aggtagaaga ggccaataaa
     5761 ggagagaaca ccagcttgtt acaccctgtg agcctgcatg ggatggatga cccggagaga
     5821 gaagtgttag agtggaggtt tgacagccgc ctagcatttc atcacgtggc ccgagagctg
     5881 catccggagt acttcaagaa ctgctgatat cgagcttgct acaagggact ttccgctggg
     5941 gactttccag ggaggcgtgg cctgggcggg actggggagt ggcgagccct cagatcctgc
     6001 atataagcag ctgctttttg cctgtactgg gtctctctgg ttagaccaga tctgagcctg
     6061 ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt
     6121 gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc
     6181 cttttagtca gtgtggaaaa tctctagcag tagtagttca tgtcatctta ttattcagta
     6241 tttataactt gcaaagaaat gaatatcaga gagtgagagg ccttgacatt gctagcgttt
     6301 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     6361 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     6421 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     6481 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     6541 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     6601 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     6661 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     6721 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     6781 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     6841 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     6901 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     6961 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     7021 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     7081 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     7141 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     7201 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     7261 ccaccgctgg tagcggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     7321 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     7381 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     7441 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     7501 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     7561 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     7621 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     7681 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     7741 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     7801 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     7861 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     7921 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     7981 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     8041 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     8101 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     8161 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     8221 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     8281 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     8341 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     8401 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     8461 gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatcaa cttgtttatt
     8521 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     8581 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg
     8641 atcaactgga taactcaagc taaccaaaat catcccaaac ttcccacccc ataccctatt
     8701 accactgcca attacctagt ggtttcattt actctaaacc tgtgattcct ctgaattatt
     8761 ttcattttaa agaaattgta tttgttaaat atgtactaca aacttagtag tttttaaaga
     8821 aattgtattt gttaaatatg tactacaaac ttagtagt
//
