LOCUS       sequence-81796-e        8163 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3rd generation lentiviral vector for enhanced eGFP 
            expression/marking in primary B cells and B-cell lines .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8163)
  AUTHORS   Mock U, Thiele R, Uhde A, Fehse B, Horn S
  TITLE     Efficient lentiviral transduction and transgene expression in 
            primary human B cells.
  JOURNAL   Hum Gene Ther Methods. 2012 Dec;23(6):408-15. doi: 
            10.1089/hgtb.2012.160.
  PUBMED    23240650
REFERENCE   2  (bases 1 to 8163)
  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: "10.1089/hgtb.2012";
            journalName: "Hum Gene Ther Methods"; date: "2012-12"; volume: "23";
            issue: "6"; pages: "408-15"
COMMENT     Sequence Label: LeGO-hEµMAR-G2
FEATURES             Location/Qualifiers
     source          1..8163
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1686..1919
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2104..2148
                     /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    2446..2563
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     primer_bind     2622..2638
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2680..2713)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        3806..4212
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     regulatory      4218..4227
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4224..4226,4227..4229,4230..4943)
                     /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(4224..4226,4227..4229,4230..4940)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4269..4290)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4530..4549)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4877..4898
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(4963..4996)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    5052..5640
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5105..5125)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5523..5534)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5643..5659)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     LTR             6169..6349
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6411..6999)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6491..6510)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7170..7961,7962..8030))
                     /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     7793..7812
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8031..8135)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg gccgcgctga tcttcagacc tggaggagga
     1561 gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca
     1621 ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg
     1681 ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcg
     1741 tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac
     1801 aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc
     1861 aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg
     1921 gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt
     1981 tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga gtgggacaga
     2041 gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca aaaccagcaa
     2101 gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg gaattggttt
     2161 aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg aggcttggta
     2221 ggtttaagaa tagtttttgc tgtactttct atagtgaata gagttaggca gggatattca
     2281 ccattatcgt ttcagaccca cctcccaacc ccgaggggac ccgacaggcc cgaaggaata
     2341 gaagaagaag gtggagagag agacagagac agatccattc gattagtgaa cggatcggca
     2401 ctgcgtgcgc caattctgca gacaaatggc agtattcatc cacaatttta aaagaaaagg
     2461 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
     2521 aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
     2581 cagcagagat ccagtttggt tagtaccggg cccgctctag tcgaggtcga cggtatcgat
     2641 aagctcgctt cacgagattc cagcaggtcg agggacctaa taacttcgta tagcatacat
     2701 tatacgaagt tatattaagg gttccaagct taagcggccg ctgaaagacc ccacctgtag
     2761 gtttggcaag ctagcaggat tgtttatctt aggaggcatg cttactgtta aaagacagga
     2821 tatgtttgaa gtggcttctg agaaaaatgg ttaagaaaat tatgacttaa aaatgtgaga
     2881 gattttcaag tatattaatt tttttaactg tccaagtatt tgaaattctt atcatttgat
     2941 taacacccat gagtgatatg tgtctggaat tgaggccaaa gcaagctcag ctaagaaata
     3001 ctagcacagt gctgtcggcc ccgatgcggg actgcgtttt gaccatcata aatcaagttt
     3061 atttttttaa ttaattgagc gaagctggaa gcagatgatg aattagagtc aagatggctg
     3121 catgggggtc tccggcaccc acagcaggtg gcaggaagca ggtcaccgcg agagtctatt
     3181 ttaggaagca aaaaaacaca attggtaaat ttatcacttc tggttgtgaa gaggtggttt
     3241 tgcccaggcc cagatctgaa agtgctctac tgagcaaaac aacacctgga caatttgcgt
     3301 ttctaaaata aggcgaggct gaccgaaact gaaaaggctt tttttaacta tctgaatttc
     3361 atttccaatc ttagcttatc aactgctagt ttgtgcaaac agcatatcaa cttctaaact
     3421 gcattcattt ttaaagtaag atgtttaaga aattaaacag tcttagggag agtttatgac
     3481 tgtattcaaa aagtttttta aattagcttg ttatcccttc atgtgataac taatctcaaa
     3541 tactttttcg atacctcaga gcattatttt cataatgact gtgttcacaa tctttttagg
     3601 ttaactcgtt ctctctttgt gattaaggag aaacactttg atattctgat agagtggcct
     3661 tcattttagt atttttcaag accacttttc aactactcac tttaggataa gttttaggta
     3721 aaatgtgcat cattatcctg aattatttca gttaagcatg ttagttggtg gcataagaga
     3781 aaactcaatc agatagctag ctgcagtaac gccattttgc aaggcatgga aaaataccaa
     3841 accaagaata gagaagttca gatcaagggc gggtacatga aaatagctaa cgttgggcca
     3901 aacaggatat ctgcggtgag cagtttcggc cccggcccgg ggccaagaac agatggtcac
     3961 cgcagtttcg gccccggccc gaggccaaga acagatggtc cccagatatg gcccaaccct
     4021 cagcagtttc ttaagaccca tcagatgttt ccaggctccc ccaaggacct gaaatgaccc
     4081 tgcgccttat ttgaattaac caatcagcct gcttctcgct tctgttcgcg cgcttctgct
     4141 tcccgagctc tataaaagag ctcacaaccc ctcactcggc gcgccagtcc tccgattgac
     4201 tgagtcgccc ggatcccgcc accatggtga gcaagggcga ggagctgttc accggggtgg
     4261 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     4321 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca
     4381 agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca
     4441 gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     4501 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     4561 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     4621 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     4681 tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg
     4741 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     4801 ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca
     4861 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     4921 gcatggacga gctgtacaag taagaattcg tcgagggacc taataacttc gtatagcata
     4981 cattatacga agttatacat gtttaagggt tccggttcca ctaggtacaa ttcgatatca
     5041 agcttatcga taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta
     5101 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta
     5161 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt
     5221 atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg
     5281 caacccccac tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt
     5341 tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag
     5401 gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc
     5461 cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc
     5521 cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc
     5581 ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc
     5641 atcgataccg tcgacctcga tcgagaccta gaaaaacatg gagcaatcac aagtagcaat
     5701 acagcagcta ccaatgctga ttgtgcctgg ctagaagcac aagaggagga ggaggtgggt
     5761 tttccagtca cacctcaggt acctttaaga ccaatgactt acaaggcagc tgtagatctt
     5821 agccactttt taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaagacaa
     5881 gatatccttg atctgtggat ctaccacaca caaggctact tccctgattg gcagaactac
     5941 acaccagggc cagggatcag atatccactg acctttggat ggtgctacaa gctagtacca
     6001 gttgagcaag agaaggtaga agaagccaat gaaggagaga acacccgctt gttacaccct
     6061 gtgagcctgc atgggatgga tgacccggag agagaagtat tagagtggag gtttgacagc
     6121 cgcctagcat ttcatcacat ggcccgagag ctgcatccgg actgtactgg gtctctctgg
     6181 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
     6241 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
     6301 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag catgtgagca
     6361 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     6421 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     6481 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     6541 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     6601 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     6661 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     6721 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     6781 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     6841 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     6901 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     6961 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     7021 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     7081 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     7141 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     7201 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     7261 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     7321 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     7381 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     7441 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     7501 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     7561 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     7621 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     7681 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     7741 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     7801 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     7861 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     7921 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     7981 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     8041 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     8101 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     8161 gac
//
