LOCUS       sequence-176751-        7365 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty backbone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7365)
  AUTHORS   Golden RJ, Chen B, Li T, Braun J, Manjunath H, Chen X, Wu J, Schmid 
            V, Chang TC, Kopp F, Ramirez-Martinez A, Tagliabracci VS, Chen ZJ, 
            Xie Y, Mendell JT
  TITLE     An Argonaute phosphorylation cycle promotes microRNA-mediated 
            silencing.
  JOURNAL   Nature. 2017 Feb 9;542(7640):197-202. doi: 10.1038/nature21025. Epub
            2017 Jan 23.
  PUBMED    28114302
REFERENCE   2  (bases 1 to 7365)
  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: 
            "10.1038/nature21025"; journalName: "Nature"; date: "2017-02-9- 9"; 
            volume: "542"; issue: "7640"; pages: "197-202"
COMMENT     Sequence Label: pLJM1-Empty
FEATURES             Location/Qualifiers
     source          1..7365
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        131..334
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     285..305
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    448..565
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        614..1124
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     1043..1061
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             1146..1745
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1146..1165)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1642..1662
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             1873..2106
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2178..2299
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2215..2234)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2269..2288
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2339..2474
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2494..2513)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2495..2513)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2523..2540)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2523..2539)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2532..2554)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2681..3136
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2768..2787)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2978..2999
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3162..3266
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3267..3335,3336..4127)
                     /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(3485..3504)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4298..4886
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4787..4806
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5040..5057
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5174..5195
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5210..5215,5216..5233,5234..5240)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5248..5264
                     /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     5253..5275
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5272..5288
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5272..5288
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5307..5327
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5309..5327
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5355..5581
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5582..5762
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5809..5934
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6427..6660
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6845..6889
                     /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"
     primer_bind     7187..7207
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
ORIGIN
        1 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
       61 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
      121 tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
      181 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
      241 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
      301 gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta
      361 gcgctaccgg gtacaagtcc ggactcagat ctcgagctca agcttcgaat tctcgacctc
      421 gagacaaatg gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag
      481 tgcaggggaa agaatagtag acataatagc aacagacata caaactaaag aattacaaaa
      541 acaaattaca aaaattcaaa attttcgggt ttattacagg gacagcagag atccactttg
      601 gccgcggctc gagggggttg gggttgcgcc ttttccaagg cagccctggg tttgcgcagg
      661 gacgcggctg ctctgggcgt ggttccggga aacgcagcgg cgccgaccct gggtctcgca
      721 cattcttcac gtccgttcgc agcgtcaccc ggatcttcgc cgctaccctt gtgggccccc
      781 cggcgacgct tcctgctccg cccctaagtc gggaaggttc cttgcggttc gcggcgtgcc
      841 ggacgtgaca aacggaagcc gcacgtctca ctagtaccct cgcagacgga cagcgccagg
      901 gagcaatggc agcgcgccga ccgcgatggg ctgtggccaa tagcggctgc tcagcagggc
      961 gcgccgagag cagcggccgg gaaggggcgg tgcgggaggc ggggtgtggg gcggtagtgt
     1021 gggccctgtt cctgcccgcg cggtgttccg cattctgcaa gcctccggag cgcacgtcgg
     1081 cagtcggctc cctcgttgac cgaatcaccg acctctctcc ccagggggat ccaccggagc
     1141 ttaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac gtccccaggg
     1201 ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac accgtcgatc
     1261 cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc
     1321 tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc
     1381 cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga
     1441 gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg caccggccca
     1501 aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag ggcaagggtc
     1561 tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg gtgcccgcct
     1621 tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc ttcaccgtca
     1681 ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc aagcccggtg
     1741 cctgacgccc gccccacgac ccgcagcgcc cgaccgaaag gagcgcacga ccccatgcat
     1801 cggtaccttt aagaccaatg acttacaagg cagctgtaga tcttagccac tttttaaaag
     1861 aaaagggggg actggaaggg ctaattcact cccaacgaag acaagatctg ctttttgctt
     1921 gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc taactaggga
     1981 acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc
     2041 tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct
     2101 ctagcagtag tagttcatgt catcttatta ttcagtattt ataacttgca aagaaatgaa
     2161 tatcagagag tgagaggaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     2221 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     2281 aactcatcaa tgtatcttat catgtctggc tctagctatc ccgcccctaa ctccgcccat
     2341 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     2401 tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg
     2461 cttttttgga ggcctaggga cgtacccaat tcgccctata gtgagtcgta ttacgcgcgc
     2521 tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat
     2581 cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat
     2641 cgcccttccc aacagttgcg cagcctgaat ggcgaatggg acgcgccctg tagcggcgca
     2701 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     2761 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     2821 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     2881 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     2941 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     3001 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     3061 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     3121 ttaacgctta caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     3181 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     3241 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     3301 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     3361 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     3421 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     3481 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     3541 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     3601 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     3661 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     3721 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     3781 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     3841 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     3901 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     3961 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     4021 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     4081 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     4141 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     4201 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     4261 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     4321 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     4381 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     4441 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     4501 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     4561 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     4621 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     4681 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     4741 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     4801 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     4861 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     4921 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     4981 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     5041 gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc
     5101 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     5161 agcgcaacgc aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta
     5221 tgcttccggc tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca
     5281 gctatgacca tgattacgcc aagcgcgcaa ttaaccctca ctaaagggaa caaaagctgg
     5341 agctgcaagc ttaatgtagt cttatgcaat actcttgtag tcttgcaaca tggtaacgat
     5401 gagttagcaa catgccttac aaggagagaa aaagcaccgt gcatgccgat tggtggaagt
     5461 aaggtggtac gatcgtgcct tattaggaag gcaacagacg ggtctgacat ggattggacg
     5521 aaccactgaa ttgccgcatt gcagagatat tgtatttaag tgcctagctc gatacataaa
     5581 cgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc
     5641 actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt
     5701 gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag
     5761 cagtggcgcc cgaacaggga cttgaaagcg aaagggaaac cagaggagct ctctcgacgc
     5821 aggactcggc ttgctgaagc gcgcacggca agaggcgagg ggcggcgact ggtgagtacg
     5881 ccaaaaattt tgactagcgg aggctagaag gagagagatg ggtgcgagag cgtcagtatt
     5941 aagcggggga gaattagatc gcgatgggaa aaaattcggt taaggccagg gggaaagaaa
     6001 aaatataaat taaaacatat agtatgggca agcagggagc tagaacgatt cgcagttaat
     6061 cctggcctgt tagaaacatc agaaggctgt agacaaatac tgggacagct acaaccatcc
     6121 cttcagacag gatcagaaga acttagatca ttatataata cagtagcaac cctctattgt
     6181 gtgcatcaaa ggatagagat aaaagacacc aaggaagctt tagacaagat agaggaagag
     6241 caaaacaaaa gtaagaccac cgcacagcaa gcggccgctg atcttcagac ctggaggagg
     6301 agatatgagg gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc
     6361 attaggagta gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt
     6421 gggaatagga gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc
     6481 gtcaatgacg ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa
     6541 caatttgctg agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat
     6601 caagcagctc caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct
     6661 ggggatttgg ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag
     6721 ttggagtaat aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag
     6781 agaaattaac aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca
     6841 agaaaagaat gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt
     6901 taacataaca aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt
     6961 aggtttaaga atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc
     7021 accattatcg tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat
     7081 agaagaagaa ggtggagaga gagacagaga cagatccatt cgattagtga acggatctcg
     7141 acggtatcga tcacgagact agcctcgagc ggccgccccc ttcaccgagg gcctatttcc
     7201 catgattcct tcatatttgc atatacgata caaggctgtt agagagataa ttggaattaa
     7261 tttgactgta aacacaaaga tattagtaca aaatacgtga cgtagaaagt aataatttct
     7321 tgggtagttt gcagttttaa aattatgttt taaaatggac tatca
//
