LOCUS       sequence-453882-        7787 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  miniEDV production plasmid. Expresses the miniGag polypeptide (CAG 
            promoter) and sgRNA (human U6 promoter).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7787)
  AUTHORS   Ngo W, Peukes J, Baldwin A, Xue ZW, Hwang S, Stickels RR, Lin Z, 
            Satpathy AT, Wells JA, Schekman R, Nogales E, Doudna JA
  TITLE     Mechanism-guided engineering of a minimal biological particle for 
            genome editing.
  JOURNAL   Proc Natl Acad Sci U S A. 2025 Jan 7;122(1):e2413519121. doi: 
            10.1073/pnas.2413519121. Epub 2024 Dec 30.
  PUBMED    39793042
REFERENCE   2  (bases 1 to 7787)
  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"; journalName: "Proc Natl 
            Acad Sci U S A."; date: "2025-01-7"; volume: "122(1):e2413519121. 
            doi"; pages: " 10.1073/pnas.2413519121. Epub 2024 Dec 30"
COMMENT     Sequence Label: pWN_U6-TRAC-miniGag
FEATURES             Location/Qualifiers
     source          1..7787
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(473..1264,1265..1333))
                     /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     1096..1115
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1334..1438)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1522..1770
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1522..1542
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1693..1712
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     enhancer        1923..2302
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2304..2581
                     /label=chicken beta-actin promoter
     intron          2582..3598
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     3522..3544
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     3606..3625
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     misc_feature    5206..5439
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             5624..5668
                     /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"
     CDS             5962..5988
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     primer_bind     complement(6273..6292)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    6338..6393
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(6392..6411)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(6754..6770)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6754..6770)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6767..6789)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6778..6794
                     /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)."
     promoter        complement(join(6802..6808,6809..6826,6827..6832))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6847..6868
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6927..7122
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      6973..7108
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    7128..7262
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7165..7184)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7219..7238
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(7330..7347)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(7501..7787,1..302))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7581..7600)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
       61 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
      121 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
      181 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
      241 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
      301 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
      361 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
      421 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
      481 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
      541 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
      601 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
      661 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
      721 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
      781 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
      841 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
      901 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
      961 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     1021 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     1081 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     1141 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     1201 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     1261 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     1321 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     1381 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     1441 catttccccg aaaagtgcca cctgggtcga ccgggtttat tacagggaca gcagagatcc
     1501 agtttggtta attaaggtac cgagggccta tttcccatga ttccttcata tttgcatata
     1561 cgatacaagg ctgttagaga gataattaga attaatttga ctgtaaacac aaagatatta
     1621 gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt tttaaaatta
     1681 tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga tttcttggct
     1741 ttatatatct tgtggaaagg acgaaacacc gcagggttct ggatatctgt gtttaagagc
     1801 tatgctggaa acagcatagc aagtttaaat aaggctagtc cgttatcaac ttgaaaaagt
     1861 ggcaccgagt cggtgctttt tttgaattcg ctagctaggt cttgaaagga gtgggaattg
     1921 tcgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
     1981 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     2041 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     2101 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
     2161 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
     2221 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
     2281 attagtcatc gctattacca tggtcgaggt gagccccacg ttctgcttca ctctccccat
     2341 ctcccccccc tccccacccc caattttgta tttatttatt ttttaattat tttgtgcagc
     2401 gatgggggcg gggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
     2461 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
     2521 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
     2581 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
     2641 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
     2701 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
     2761 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg
     2821 cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg
     2881 ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg
     2941 gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt
     3001 gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca
     3061 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     3121 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc
     3181 ggggccgcct cgggccgggg agggctcggg ggaggggcgc ggcggccccc ggagcgccgg
     3241 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     3301 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     3361 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     3421 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     3481 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     3541 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     3601 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tcgggccgcg
     3661 ttgacgcgca cggcaagagg cgaggggcgg cgactggtga gagatgggtg cgagagcgtc
     3721 agtattaagc gggggagaat tagatcgatg ggaaaaaatt cggttaaggc cagggggaaa
     3781 gaaaaaatat aaattaaaac atatagtatg ggcaagcagg gagctagaac gattcgcagt
     3841 taatcctggc ctgttagaaa catcagaagg ctgtagacaa atactgggac agctacaacc
     3901 atcccttcag acaggatcag aagaacttag atcattatat aatacagtag caaccctcta
     3961 ttgtgtgcat caaaggatag agataaaaga caccaaggaa gctttagaca agatagagga
     4021 agagcaaaac aaaagtaaga aaaaagcaca gcaagcagca gctgacacag gacacagcaa
     4081 tcaggtcagc caaaattacc ctatagtgca gagcattctg gacataagac aaggaccaaa
     4141 ggaacccttt agagactatg tagaccgatt ctataaaact ctaagagccg agcaagcttc
     4201 acaagaggta aaaaattgga tgacagaaac cttgttggtc caaaatgcga acccagattg
     4261 taagactatt ttaaaagcat tgggaccagg agcgacacta gaagaaatga tgacagcatg
     4321 tcagggagtg gggggacccg gccataaagc aagagttttg gctgaagcaa tgagccaagt
     4381 aacaaatcca gctaccataa tgatacagaa aggcagacag gctaattttt tagggaagat
     4441 ctggccttcc cacaagggaa ggccagggaa ttttcttcag agcagaccag agccaacagc
     4501 cccaccagaa gagagcttca ggtttgggga agagacaaca actccctctc agaagcagga
     4561 gccgatagac aaggaactgt atcctttagc ttccctcaga tcactctttg gcagcgaccc
     4621 ctcgtcacaa taacacatgg aattctgcaa caactgctgt ttatccattt cagaattggg
     4681 ttcgacatag cagaataggc gttactcgac agaggagagc aagaaatgga gccagtagat
     4741 cctagactag agccctggaa gcatccagga agtcagccta aaactgcttg taccaattgc
     4801 tattgtaaaa agtgttgctt tcattgccaa gtttgtttca tgacaaaagc cttaggcatc
     4861 tcctatggca ggaagaagcg gagacagcga cgaagagctc atcagaacag tcagactcat
     4921 caagcttctc tatcaaagca gtaagtagta catgtaatgc aacctataat agtagcaata
     4981 gtagcattag tagtagcaat aataatagca atagttgtgt ggtccatagt aatcatagaa
     5041 tataggaaaa tggccgctga tcttcagacc tggaggagga gatatgaggg acaattggag
     5101 aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag cacccaccaa
     5161 ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag ctttgttcct
     5221 tgggttcttg ggagcagcag gaagcactat gggcgcagcg tcaatgacgc tgacggtaca
     5281 ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga gggctattga
     5341 ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc aggcaagaat
     5401 cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg gttgctctgg
     5461 aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata aatctctgga
     5521 acagatttgg aatcacacga cctggatgga gtgggacaga gaaattaaca attacacaag
     5581 cttaatacac tccttaattg aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt
     5641 attggaatta gataaatggg caagtttgtg gaattggttt aacataacaa attggctgtg
     5701 gtatataaaa ttattcataa tgatagtagg aggcttggta ggtttaagaa tagtttttgc
     5761 tgtactttct atagtgaata gagttaggca gggatattca ccattatcgt ttcagaccca
     5821 cctcccaacc ccgaggggac ccgacaggcc cgaaggaata gaagaagaag gtggagagag
     5881 agacagagac agatccattc gattagtgaa cggatccttg gcacttatct gggacgatct
     5941 gcggagcctg tgcctcttca gctaccaccg cttgagagac ttactcttga ttgtaacgag
     6001 gattgtggaa cttctgggac gcagggggtg ggaagccctc aaatattggt ggaatctcct
     6061 acaatattgg agtcaggagc taaagaatag tgctgttagc ttgctcaatg ccacagccat
     6121 agcagtagct gaggggacag atagggttat agaagtagta caaggagctt gtagagctat
     6181 tcgccacata cctagaagaa taagacaggg cttggaaagg attttgctat aagctcgaaa
     6241 caaccggtac ctctagaact atagctagca gatctttttc cctctgccaa aaattatggg
     6301 gacatcatga agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt
     6361 gcaatagtgt gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat
     6421 catttaaaac atcagaatga gtatttggtt tagagtttgg caacatatgc ccatatgctg
     6481 gctgccatga acaaaggttg gctataaaga ggtcatcagt atatgaaaca gccccctgct
     6541 gtccattcct tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt
     6601 tttgtgttat ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga
     6661 tttttcctcc tctcctgact actcccagtc atagctgtcc ctcttctctt atggagatcc
     6721 ctcgacctgc agcccaagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     6781 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     6841 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     6901 gggaaacctg tcgtgccagc ggatccgcat ctcaattagt cagcaaccat agtcccgccc
     6961 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     7021 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     7081 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctaac ttgtttattg
     7141 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     7201 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga
     7261 tccgctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     7321 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     7381 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     7441 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     7501 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     7561 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     7621 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     7681 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     7741 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcg
//
