LOCUS Exported 8282 bp ds-DNA circular SYN 08-JAN-2024 DEFINITION 3rd gen lentiviral eGFP expression vector, PGK promoter, Blast. ACCESSION . VERSION . KEYWORDS pLenti PGK GFP Blast (w510-5) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8282) AUTHORS Campeau E, Ruhl VE, Rodier F, Smith CL, Rahmberg BL, Fuss JO, Campisi J, Yaswen P, Cooper PK, Kaufman PD. TITLE A versatile viral system for expression and depletion of proteins in mammalian cells. JOURNAL PLoS One. 2009 Aug 6;4(8):e6529. PUBMED 19657394 REFERENCE 2 (bases 1 to 8282) AUTHORS . TITLE Direct Submission JOURNAL Exported Jan 8, 2024 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8282 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 27..131 /gene="bla" /label=AmpR promoter CDS 132..992 /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 complement(350..369) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 1163..1751 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 1652..1671 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 1905..1922 /label=L4440 /note="L4440 vector, forward primer" protein_bind 2039..2060 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 2075..2105 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 2113..2129 /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)." primer_bind 2118..2140 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 2137..2153 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 2137..2153 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind 2172..2192 /label=T3 /note="T3 promoter, forward primer" promoter 2174..2192 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" LTR 2448..2628 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 2672..2797 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 3290..3523 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 3708..3752 /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 4019..4136 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 4183..4693 /label=hPGK promoter /note="human phosphoglycerate kinase 1 promoter" primer_bind 4612..4630 /label=hPGK-F /note="Human PGK promoter, forward primer" regulatory 4709..4718 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 4715..5434 /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 4715..5431 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(4760..4781) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(5021..5040) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 5368..5389 /label=EGFP-C /note="EGFP, forward primer" misc_feature 5450..6038 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(5503..5523) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(5921..5932) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(6089..6109) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 6094..6424 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 6275..6410 /label=SV40 ori /note="SV40 origin of replication" promoter 6472..6519 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 6538..6936 /codon_start=1 /gene="Aspergillus terreus BSD" /product="blasticidin S deaminase" /label=BSD /note="confers resistance to blasticidin" /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI KAIVKDSDGQPTAVGIRELLPSGYVWEG" LTR 7023..7256 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" polyA_signal 7328..7449 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(7365..7384) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 7419..7438 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin 7468..7603 /label=SV40 ori /note="SV40 origin of replication" primer_bind complement(7635..7654) /label=T7 /note="T7 promoter, forward primer" promoter complement(7636..7654) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(7664..7681) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(7664..7680) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(7673..7695) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 7825..8280 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(7912..7931) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 8122..8143 /label=F1ori-F /note="F1 origin, forward primer" ORIGIN 1 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac 61 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa 121 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat 181 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc 241 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga 301 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg 361 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc 421 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag 481 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc 541 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg 601 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg 661 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac 721 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac 781 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg 841 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg 901 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg 961 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac 1021 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg 1081 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg 1141 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc 1201 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc 1261 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt 1321 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc 1381 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact 1441 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac 1501 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag 1561 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg 1621 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg 1681 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga 1741 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt 1801 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct 1861 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg 1921 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt 1981 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta 2041 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta 2101 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt 2161 acgccaagcg cgcaattaac cctcactaaa gggaacaaaa gctggagctg caagcttaat 2221 gtagtcttat gcaatactct tgtagtcttg caacatggta acgatgagtt agcaacatgc 2281 cttacaagga gagaaaaagc accgtgcatg ccgattggtg gaagtaaggt ggtacgatcg 2341 tgccttatta ggaaggcaac agacgggtct gacatggatt ggacgaacca ctgaattgcc 2401 gcattgcaga gatattgtat ttaagtgcct agctcgatac aataaacggg tctctctggt 2461 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc 2521 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta 2581 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa 2641 cagggacctg aaagcgaaag ggaaaccaga gctctctcga cgcaggactc ggcttgctga 2701 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag 2761 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag 2821 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca 2881 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac 2941 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga 3001 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga 3061 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac 3121 caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt 3181 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca 3241 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt 3301 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcctcaatg acgctgacgg 3361 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta 3421 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa 3481 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct 3541 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc 3601 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca 3661 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag 3721 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc 3781 tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt 3841 ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga 3901 cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag 3961 agagagacag agacagatcc attcgattag tgaacggatc tcgacggtat cggttaactt 4021 ttaaaagaaa aggggggatt ggggggtaca gtgcagggga aagaatagta gacataatag 4081 caacagacat acaaactaaa gaattacaaa aacaaattac aaaaattcaa aattttatcg 4141 atcacgagac tagcctcgag aagcttgata tcgaattccc acggggttgg ggttgcgcct 4201 tttccaaggc agccctgggt ttgcgcaggg acgcggctgc tctgggcgtg gttccgggaa 4261 acgcagcggc gccgaccctg ggtctcgcac attcttcacg tccgttcgca gcgtcacccg 4321 gatcttcgcc gctacccttg tgggcccccc ggcgacgctt cctgctccgc ccctaagtcg 4381 ggaaggttcc ttgcggttcg cggcgtgccg gacgtgacaa acggaagccg cacgtctcac 4441 tagtaccctc gcagacggac agcgccaggg agcaatggca gcgcgccgac cgcgatgggc 4501 tgtggccaat agcggctgct cagcggggcg cgccgagagc agcggccggg aaggggcggt 4561 gcgggaggcg gggtgtgggg cggtagtgtg ggccctgttc ctgcccgcgc ggtgttccgc 4621 attctgcaag cctccggagc gcacgtcggc agtcggctcc ctcgttgacc gaatcaccga 4681 cctctctccc cagggggatc caccggtcgc caccatggtg agcaagggcg aggagctgtt 4741 caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag 4801 cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg 4861 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt 4921 gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat 4981 gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac 5041 ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat 5101 cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca 5161 caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg 5221 ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat 5281 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag 5341 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg 5401 gatcactctc ggcatggacg agctgtacaa gtaaagcggc cgcgtcgaca atcaacctct 5461 ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct 5521 atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat 5581 tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt 5641 caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat 5701 tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc 5761 ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga 5821 caattccgtg gtgttgtcgg ggaagctgac gtcctttcca tggctgctcg cctgtgttgc 5881 cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga 5941 ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc 6001 tcagacgagt cggatctccc tttgggccgc ctccccgcct ggaattctgc agatatccgg 6061 ttagtaatga gtttggaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc 6121 ccaggctccc caggcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag 6181 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta 6241 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc 6301 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc 6361 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg 6421 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg tgttgacaat 6481 taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa ctaaaccatg 6541 gccaagcctt tgtctcaaga agaatccacc ctcattgaaa gagcaacggc tacaatcaac 6601 agcatcccca tctctgaaga ctacagcgtc gccagcgcag ctctctctag cgacggccgc 6661 atcttcactg gtgtcaatgt atatcatttt actgggggac cttgtgcaga actcgtggtg 6721 ctgggcactg ctgctgctgc ggcagctggc aacctgactt gtatcgtcgc gatcggaaat 6781 gagaacaggg gcatcttgag cccctgcgga cggtgccgac aggtgcttct cgatctgcat 6841 cctgggatca aagccatagt gaaggacagt gatggacagc cgacggcagt tgggattcgt 6901 gaattgctgc cctctggtta tgtgtgggag ggctaagcac aattcgagct cggtaccttt 6961 aagaccaatg acttacaagg cagctgtaga tcttagccac tttttaaaag aaaagggggg 7021 actggaaggg ctagctcact cccaacgaag acaagatctg ctttttgctt gtactgggtc 7081 tctctggtta gaccagatct gagcctggga gctctctggc tgcctaggga acccactgct 7141 taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga 7201 ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtag 7261 tagttcatgt catcttatta ttcagtattt ataacttgca aagaaatgaa tatcagagag 7321 tgagaggaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa 7381 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa 7441 tgtatcttat catgtctggc tctagctatc ccgcccctaa ctccgcccag ttccgcccat 7501 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc 7561 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcgtcgag 7621 acgtacccaa ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac 7681 aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc 7741 ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc 7801 gcagcctgaa tggcgaatgg cgcgacgcgc cctgtagcgg cgcattaagc gcggcgggtg 7861 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg 7921 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg 7981 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt 8041 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt 8101 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta 8161 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa 8221 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt 8281 cc //