LOCUS       sequence-220221-        8084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses mEGFP-TSG101 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8084)
  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 8084)
  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: 
            "10.1073/pnas.1321655111"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2014-08-19- 19"; volume: "111"; issue: "33"; pages: "12211-6"
COMMENT     Sequence Label: pLNCX2-mEGFP-TSG101
FEATURES             Location/Qualifiers
     source          1..8084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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        2378..2681
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2682..2885
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2836..2856
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2930..2949
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2930..2948
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      2976..2985
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2982..2984,2985..2987,2988..3698)
                     /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(3027..3048)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3288..3307)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3635..3656
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             4986..5579
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5682..5704)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5790..5930
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5845..5864
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5945..5962)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6116..6704)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6196..6215)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6875..7666,7667..7735))
                     /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     7498..7517
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7736..7840)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7908..7926
                     /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 ttcgtaatag taatcaatta
     2341 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
     2401 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
     2461 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
     2521 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
     2581 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
     2641 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
     2701 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
     2761 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
     2821 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
     2881 gagctctctg gctaactaga gaacccactg cttactggct tatcgaaatt aatacgactc
     2941 actataggga gacccaagct ctagcgctac cggtcgccac catggtgagc aagggcgagg
     3001 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     3061 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
     3121 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct
     3181 acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     3241 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     3301 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     3361 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     3421 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca
     3481 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     3541 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtcca
     3601 agctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     3661 ccgccgggat cactctcggc atggacgagc tgtacaagtc cggactcaga tccccggaat
     3721 tcatggcggt gtcggagagc cagctcaaga aaatggtgtc caagtacaaa tacagagacc
     3781 taactgtacg tgaaactgtc aatgttatta ctctatacaa agatctcaaa cctgttttgg
     3841 attcatatgt ttttaacgat ggcagttcca gggaactaat gaacctcact ggaacaatcc
     3901 ctgtgcctta tagaggtaat acatacaata ttccaatatg cctatggcta ctggacacat
     3961 acccatataa tccccctatc tgttttgtta agcctactag ttcaatgact attaaaacag
     4021 gaaagcatgt tgatgcaaat gggaagatat atcttcctta tctacatgaa tggaaacacc
     4081 cacagtcaga cttgttgggg cttattcagg tcatgattgt ggtatttgga gatgaacctc
     4141 cagtcttctc tcgtcctatt tcggcatcct atccgccata ccaggcaacg gggccaccaa
     4201 atacttccta catgccaggc atgccaggtg gaatctctcc atacccatcc ggataccctc
     4261 ccaatcccag tggttaccca ggctgtcctt acccacctgg tggtccatat cctgccacaa
     4321 caagttctca gtacccttct cagcctcctg tgaccactgt tggtcccagt agggatggca
     4381 caatcagcga ggacaccatc cgagcctctc tcatctctgc ggtcagtgac aaactgagat
     4441 ggcggatgaa ggaggaaatg gatcgtgccc aggcagagct caatgccttg aaacgaacag
     4501 aagaagacct gaaaaagggt caccagaaac tggaagagat ggttacccgt ttagatcaag
     4561 aagtagccga ggttgataaa aacatagaac ttttgaaaaa gaaggatgaa gaactcagtt
     4621 ctgctctgga aaaaatggaa aatcagtctg aaaacaatga tatcgatgaa gttatcattc
     4681 ccacagctcc cttatacaaa cagatcctga atctgtatgc agaagaaaac gctattgaag
     4741 acactatctt ttacttggga gaagccttga gaaggggcgt gatagacctg gatgtcttcc
     4801 tgaagcatgt acgtcttctg tcccgtaaac agttccagct gagggcacta atgcaaaaag
     4861 caagaaagac tgccggtctc agtgacctct actgactcga cctgcagcgg ccgcttgtcg
     4921 acaggcctta atggcctaac atcgataaaa taaaagattt tatttagtct ccagaaaaag
     4981 gggggaatga aagaccccac ctgtaggttt ggcaagctag cttaagtaac gccattttgc
     5041 aaggcatgga aaaatacata actgagaata gagaagttca gatcaaggtc aggaacagat
     5101 ggaacagctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg ccccggctca
     5161 gggccaagaa cagatggaac agctgaatat gggccaaaca ggatatctgt ggtaagcagt
     5221 tcctgccccg gctcagggcc aagaacagat ggtccccaga tgcggtccag ccctcagcag
     5281 tttctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg accctgtgcc
     5341 ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc tgctccccga
     5401 gctcaataaa agagcccaca acccctcact cggggcgcca gtcctccgat tgactgagtc
     5461 gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt gtggtctcgc
     5521 tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg tctttcattt
     5581 gggggctcgt ccgggatcgg gagacccctg cccagggacc accgacccac caccgggagg
     5641 taagctggct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc
     5701 ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc
     5761 gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg tagcgatagc
     5821 ggagtgtata ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata
     5881 tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgctcttccg
     5941 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     6001 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     6061 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     6121 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     6181 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     6241 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     6301 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     6361 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     6421 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     6481 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     6541 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     6601 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     6661 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     6721 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     6781 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     6841 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     6901 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     6961 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     7021 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     7081 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     7141 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct gcaggcatcg
     7201 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     7261 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     7321 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     7381 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     7441 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca acacgggata
     7501 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     7561 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     7621 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     7681 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     7741 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     7801 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     7861 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
     7921 cgaggccctt tcgtcttcaa gaattcatac cagatcaccg aaaactgtcc tccaaatgtg
     7981 tccccctcac actcccaaat tcgcgggctt ctgcctctta gaccactcta ccctattccc
     8041 cacactcacc ggagccaaag ccgcggccct tccgtttctt tgct
//
