LOCUS       sequence-223905-        7567 bp    DNA     circular SYN 14-APR-2025
DEFINITION  expresses mEGFP-CHMP2B in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7567)
  AUTHORS   Johnson DS, Bleck M, Simon SM
  TITLE     Timing of ESCRT-III protein recruitment and membrane scission during
            HIV-1 assembly.
  JOURNAL   Elife. 2018 Jul 4;7. pii: 36221. doi: 10.7554/eLife.36221.
  PUBMED    29972351
REFERENCE   2  (bases 1 to 7567)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Elife."; 
            date: "2018-07-4"; volume: "7. pii: 36221. doi"; pages: " 
            10.7554/eLife.36221"
COMMENT     Sequence Label: pLNCX2-mEGFP-CHMP2B
FEATURES             Location/Qualifiers
     source          1..7567
                     /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        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
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2981..2983,2984..2986,2987..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             4469..5062
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5165..5187)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5273..5413
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5328..5347
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5428..5445)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5599..6187)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5679..5698)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6358..7149,7150..7218))
                     /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     6981..7000
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7219..7323)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7391..7409
                     /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 catggcgtcc ctcttcaaga agaaaaccgt ggatgatgta ataaaggaac agaatcgaga
     3781 gttacgaggt acacagaggg ctataatcag agatcgagca gctttagaga aacaagaaaa
     3841 acagctggaa ttagaaatta agaaaatggc caagattggt aataaggaag cttgcaaagt
     3901 tttagccaaa caacttgtgc atctacggaa acagaagacg agaacttttg ctgtaagttc
     3961 aaaagttact tctatgtcta cacaaacaaa agtgatgaat tcccaaatga agatggctgg
     4021 agcaatgtct accacagcaa aaacaatgca ggcagttaac aagaagatgg atccacaaaa
     4081 gacattacaa acaatgcaga atttccagaa ggaaaacatg aaaatggaaa tgactgaaga
     4141 aatgatcaat gatacacttg atgacatctt tgacggttct gatgacgaag aagaaagcca
     4201 ggatattgtg aatcaagttc ttgatgaaat tggaattgaa atttctggaa agatggccaa
     4261 agctccatca gctgctcgaa gcttaccatc tgcctctact tcaaaggcta caatctcaga
     4321 tgaagagatt gaacggcaac tcaaggcttt aggagtagat tagctcgagc tcaagcttgt
     4381 ttggccgagg cggccgcttg tcgacaggcc ttaatggcct aacatcgata aaataaaaga
     4441 ttttatttag tctccagaaa aaggggggaa tgaaagaccc cacctgtagg tttggcaagc
     4501 tagcttaagt aacgccattt tgcaaggcat ggaaaaatac ataactgaga atagagaagt
     4561 tcagatcaag gtcaggaaca gatggaacag ctgaatatgg gccaaacagg atatctgtgg
     4621 taagcagttc ctgccccggc tcagggccaa gaacagatgg aacagctgaa tatgggccaa
     4681 acaggatatc tgtggtaagc agttcctgcc ccggctcagg gccaagaaca gatggtcccc
     4741 agatgcggtc cagccctcag cagtttctag agaaccatca gatgtttcca gggtgcccca
     4801 aggacctgaa atgaccctgt gccttatttg aactaaccaa tcagttcgct tctcgcttct
     4861 gttcgcgcgc ttctgctccc cgagctcaat aaaagagccc acaacccctc actcggggcg
     4921 ccagtcctcc gattgactga gtcgcccggg tacccgtgta tccaataaac cctcttgcag
     4981 ttgcatccga cttgtggtct cgctgttcct tgggagggtc tcctctgagt gattgactac
     5041 ccgtcagcgg gggtctttca tttgggggct cgtccgggat cgggagaccc ctgcccaggg
     5101 accaccgacc caccaccggg aggtaagctg gctgcctcgc gcgtttcggt gatgacggtg
     5161 aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg
     5221 ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca
     5281 tgacccagtc acgtagcgat agcggagtgt atactggctt aactatgcgg catcagagca
     5341 gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg taaggagaaa
     5401 ataccgcatc aggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     5461 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     5521 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     5581 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     5641 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     5701 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     5761 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     5821 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     5881 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     5941 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     6001 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     6061 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     6121 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     6181 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     6241 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     6301 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     6361 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     6421 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     6481 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     6541 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     6601 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     6661 tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     6721 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     6781 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     6841 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     6901 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     6961 ttgcccggcg tcaacacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     7021 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     7081 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     7141 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     7201 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     7261 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     7321 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
     7381 aacctataaa aataggcgta tcacgaggcc ctttcgtctt caagaattca taccagatca
     7441 ccgaaaactg tcctccaaat gtgtccccct cacactccca aattcgcggg cttctgcctc
     7501 ttagaccact ctaccctatt ccccacactc accggagcca aagccgcggc ccttccgttt
     7561 ctttgct
//
