LOCUS Exported 8083 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION expresses mEGFP-TSG101 in mammalian cells. ACCESSION . VERSION . KEYWORDS pLNCX2-mEGFP-TSG101 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8083) AUTHORS Bleck M, Itano MS, Johnson DS, Thomas VK, North AJ, Bieniasz PD, Simon SM TITLE Temporal and spatial organization of ESCRT protein recruitment during HIV-1 budding. JOURNAL Proc Natl Acad Sci U S A. 2014 Aug 19;111(33):12211-6. doi: 10.1073/pnas.1321655111. Epub 2014 Aug 6. PUBMED 25099357 REFERENCE 2 (bases 1 to 8083) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8083 /organism="synthetic DNA construct" /mol_type="other DNA" LTR 414..589 /label=5' LTR (truncated) /note="truncated long terminal repeat from Moloney murine sarcoma virus" misc_feature 652..851 /label=MMLV Psi /note="packaging signal of Moloney murine leukemia virus (MMLV)" CDS 1052..1468 /label=gag (truncated) /note="truncated Moloney murine leukemia virus (MMLV) gag gene lacking the start codon" primer_bind 1398..1420 /label=pLXSN 5' /note="Murine stem cell virus, forward primer. Also called MSCV" primer_bind 1436..1452 /label=pBABE 5' /note="Psi packaging signal, forward primer for pBABE vectors" CDS 1512..2306 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(1566..1585) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 2176..2195 /label=Neo-F /note="Neomycin resistance gene, forward primer" enhancer 2377..2680 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 2681..2884 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 2835..2855 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 2929..2948 /label=T7 /note="T7 promoter, forward primer" promoter 2929..2947 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" regulatory 2975..2984 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2981..3697 /codon_start=1 /product="enhanced GFP with monomerizing A206K mutation (Zacharias et al., 2002)" /label=mEGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(3026..3047) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(3287..3306) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 3634..3655 /label=EGFP-C /note="EGFP, forward primer" LTR 4985..5578 /note="long terminal repeat from Moloney murine leukemia virus" primer_bind complement(5681..5703) /label=pGEX 3' /note="pGEX vectors, reverse primer" misc_feature 5789..5929 /label=bom /note="basis of mobility region from pBR322" primer_bind 5844..5863 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind complement(5944..5961) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(6115..6703) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(6195..6214) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6874..7734) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind 7497..7516 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7735..7839) /gene="bla" /label=AmpR promoter primer_bind 7907..7925 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" ORIGIN 1 tttgaaagac cccacccgta ggtggcaagc tagcttaagt aacgccactt tgcaaggcat 61 ggaaaaatac ataactgaga atagaaaagt tcagatcaag gtcaggaaca aagaaacagc 121 tgaataccaa acaggatatc tgtggtaagc ggttcctgcc ccggctcagg gccaagaaca 181 gatgagacag ctgagtgatg ggccaaacag gatatctgtg gtaagcagtt cctgccccgg 241 ctcggggcca agaacagatg gtccccagat gcggtccagc cctcagcagt ttctagtgaa 301 tcatcagatg tttccagggt gccccaagga cctgaaaatg accctgtacc ttatttgaac 361 taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc cgctctccga gctcaataaa 421 agagcccaca acccctcact cggcgcgcca gtcttccgat agactgcgtc gcccgggtac 481 ccgtattccc aataaagcct cttgctgttt gcatccgaat cgtggtctcg ctgttccttg 541 ggagggtctc ctctgagtga ttgactaccc acgacggggg tctttcattt gggggctcgt 601 ccgggatttg gagacccctg cccagggacc accgacccac caccgggagg taagctggcc 661 agcaacttat ctgtgtctgt ccgattgtct agtgtctatg tttgatgtta tgcgcctgcg 721 tctgtactag ttagctaact agctctgtat ctggcggacc cgtggtggaa ctgacgagtt 781 ctgaacaccc ggccgcaacc ctgggagacg tcccagggac tttgggggcc gtttttgtgg 841 cccgacctga ggaagggagt cgatgtggaa tccgaccccg tcaggatatg tggttctggt 901 aggagacgag aacctaaaac agttcccgcc tccgtctgaa tttttgcttt cggtttggaa 961 ccgaagccgc gcgtcttgtc tgctgcagcg ctgcagcatc gttctgtgtt gtctctgtct 1021 gactgtgttt ctgtatttgt ctgaaaatta gggccagact gttaccactc ccttaagttt 1081 gaccttaggt cactggaaag atgtcgagcg gatcgctcac aaccagtcgg tagatgtcaa 1141 gaagagacgt tgggttacct tctgctctgc agaatggcca acctttaacg tcggatggcc 1201 gcgagacggc acctttaacc gagacctcat cacccaggtt aagatcaagg tcttttcacc 1261 tggcccgcat ggacacccag accaggtccc ctacatcgtg acctgggaag ccttggcttt 1321 tgacccccct ccctgggtca agccctttgt acaccctaag cctccgcctc ctcttcctcc 1381 atccgccccg tctctccccc ttgaacctcc tcgttcgacc ccgcctcgat cctcccttta 1441 tccagccctc actccttctc taggcgccgg aattccgatc tgatcaagag acaggatgag 1501 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg 1561 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt 1621 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc 1681 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt 1741 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag 1801 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg 1861 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag 1921 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg 1981 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc 2041 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca 2101 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc 2161 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg 2221 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct 2281 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaatagt aatcaattac 2341 ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg 2401 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc 2461 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac 2521 tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa 2581 tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac 2641 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta 2701 catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga 2761 cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa 2821 ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag 2881 agctctctgg ctaactagag aacccactgc ttactggctt atcgaaatta atacgactca 2941 ctatagggag acccaagctc tagcgctacc ggtcgccacc atggtgagca agggcgagga 3001 gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa 3061 gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt 3121 catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta 3181 cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc 3241 cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta 3301 caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa 3361 gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa 3421 cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa 3481 gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac 3541 ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccaa 3601 gctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc 3661 cgccgggatc actctcggca tggacgagct gtacaagtcc ggactcagat ccccggaatt 3721 catggcggtg tcggagagcc agctcaagaa aatggtgtcc aagtacaaat acagagacct 3781 aactgtacgt gaaactgtca atgttattac tctatacaaa gatctcaaac ctgttttgga 3841 ttcatatgtt tttaacgatg gcagttccag ggaactaatg aacctcactg gaacaatccc 3901 tgtgccttat agaggtaata catacaatat tccaatatgc ctatggctac tggacacata 3961 cccatataat ccccctatct gttttgttaa gcctactagt tcaatgacta ttaaaacagg 4021 aaagcatgtt gatgcaaatg ggaagatata tcttccttat ctacatgaat ggaaacaccc 4081 acagtcagac ttgttggggc ttattcaggt catgattgtg gtatttggag atgaacctcc 4141 agtcttctct cgtcctattt cggcatccta tccgccatac caggcaacgg ggccaccaaa 4201 tacttcctac atgccaggca tgccaggtgg aatctctcca tacccatccg gataccctcc 4261 caatcccagt ggttacccag gctgtcctta cccacctggt ggtccatatc ctgccacaac 4321 aagttctcag tacccttctc agcctcctgt gaccactgtt ggtcccagta gggatggcac 4381 aatcagcgag gacaccatcc gagcctctct catctctgcg gtcagtgaca aactgagatg 4441 gcggatgaag gaggaaatgg atcgtgccca ggcagagctc aatgccttga aacgaacaga 4501 agaagacctg aaaaagggtc accagaaact ggaagagatg gttacccgtt tagatcaaga 4561 agtagccgag gttgataaaa acatagaact tttgaaaaag aaggatgaag aactcagttc 4621 tgctctggaa aaaatggaaa atcagtctga aaacaatgat atcgatgaag ttatcattcc 4681 cacagctccc ttatacaaac agatcctgaa tctgtatgca gaagaaaacg ctattgaaga 4741 cactatcttt tacttgggag aagccttgag aaggggcgtg atagacctgg atgtcttcct 4801 gaagcatgta cgtcttctgt cccgtaaaca gttccagctg agggcactaa tgcaaaaagc 4861 aagaaagact gccggtctca gtgacctcta ctgactcgac ctgcagcggc cgcttgtcga 4921 caggccttaa tggcctaaca tcgataaaat aaaagatttt atttagtctc cagaaaaagg 4981 ggggaatgaa agaccccacc tgtaggtttg gcaagctagc ttaagtaacg ccattttgca 5041 aggcatggaa aaatacataa ctgagaatag agaagttcag atcaaggtca ggaacagatg 5101 gaacagctga atatgggcca aacaggatat ctgtggtaag cagttcctgc cccggctcag 5161 ggccaagaac agatggaaca gctgaatatg ggccaaacag gatatctgtg gtaagcagtt 5221 cctgccccgg ctcagggcca agaacagatg gtccccagat gcggtccagc cctcagcagt 5281 ttctagagaa ccatcagatg tttccagggt gccccaagga cctgaaatga ccctgtgcct 5341 tatttgaact aaccaatcag ttcgcttctc gcttctgttc gcgcgcttct gctccccgag 5401 ctcaataaaa gagcccacaa cccctcactc ggggcgccag tcctccgatt gactgagtcg 5461 cccgggtacc cgtgtatcca ataaaccctc ttgcagttgc atccgacttg tggtctcgct 5521 gttccttggg agggtctcct ctgagtgatt gactacccgt cagcgggggt ctttcatttg 5581 ggggctcgtc cgggatcggg agacccctgc ccagggacca ccgacccacc accgggaggt 5641 aagctggctg cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc 5701 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg 5761 cgtcagcggg tgttggcggg tgtcggggcg cagccatgac ccagtcacgt agcgatagcg 5821 gagtgtatac tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat 5881 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gctcttccgc 5941 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca 6001 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg 6061 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca 6121 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa 6181 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc 6241 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc 6301 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct 6361 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg 6421 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag 6481 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta 6541 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg 6601 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt 6661 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt 6721 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag 6781 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat 6841 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc 6901 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat 6961 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc 7021 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag 7081 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag 7141 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgctg caggcatcgt 7201 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg 7261 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt 7321 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc 7381 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc 7441 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa cacgggataa 7501 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg 7561 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc 7621 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag 7681 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt 7741 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt 7801 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc 7861 acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac 7921 gaggcccttt cgtcttcaag aattcatacc agatcaccga aaactgtcct ccaaatgtgt 7981 ccccctcaca ctcccaaatt cgcgggcttc tgcctcttag accactctac cctattcccc 8041 acactcaccg gagccaaagc cgcggccctt ccgtttcttt gct //