LOCUS       sequence-269251-        7084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SMG6 shRNA (Targeting CDS #2) inserted into the PLKO.1 plasmid 
            (AAGGAGTTCCAGGTGTTACTG).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7084)
  AUTHORS   Fischer JW, Busa VF, Shao Y, Leung AKL
  TITLE     Structure-Mediated RNA Decay by UPF1 and G3BP1.
  JOURNAL   Mol Cell. 2020 Apr 2;78(1):70-84.e6. doi: 
            10.1016/j.molcel.2020.01.021. Epub 2020 Feb 3.
  PUBMED    32017897
REFERENCE   2  (bases 1 to 7084)
  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: 
            "10.1016/j.molcel.2020.01.021"; journalName: "Mol Cell"; date: 
            "2020-04-2- 2"; volume: "78"; issue: "1"; pages: "70-84.e6"
COMMENT     Sequence Label: PLKO.1-SMG6-CDS-2
FEATURES             Location/Qualifiers
     source          1..7084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             85..129
                     /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"
     promoter        427..675
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     427..447
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     598..617
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    772..889
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        938..1448
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     1367..1385
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             1470..2069
                     /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(1470..1489)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1966..1986
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             2197..2430
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2502..2623
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2539..2558)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2593..2612
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2663..2798
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2818..2837)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2819..2837)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2847..2864)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2847..2863)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2856..2878)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3005..3460
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3092..3111)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3302..3323
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3486..3590
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3591..3659,3660..4451)
                     /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(3809..3828)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4622..5210
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5111..5130
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5364..5381
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5498..5519
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5534..5539,5540..5557,5558..5564)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5572..5588
                     /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     5577..5599
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5596..5612
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5596..5612
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5631..5651
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5633..5651
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5679..5905
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5906..6086
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6133..6258
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6751..6984
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
ORIGIN
        1 acctggatgg agtgggacag agaaattaac aattacacaa gcttaataca ctccttaatt
       61 gaagaatcgc aaaaccagca agaaaagaat gaacaagaat tattggaatt agataaatgg
      121 gcaagtttgt ggaattggtt taacataaca aattggctgt ggtatataaa attattcata
      181 atgatagtag gaggcttggt aggtttaaga atagtttttg ctgtactttc tatagtgaat
      241 agagttaggc agggatattc accattatcg tttcagaccc acctcccaac cccgagggga
      301 cccgacaggc ccgaaggaat agaagaagaa ggtggagaga gagacagaga cagatccatt
      361 cgattagtga acggatctcg acggtatcga tcacgagact agcctcgagc ggccgccccc
      421 ttcaccgagg gcctatttcc catgattcct tcatatttgc atatacgata caaggctgtt
      481 agagagataa ttggaattaa tttgactgta aacacaaaga tattagtaca aaatacgtga
      541 cgtagaaagt aataatttct tgggtagttt gcagttttaa aattatgttt taaaatggac
      601 tatcatatgc ttaccgtaac ttgaaagtat ttcgatttct tggctttata tatcttgtgg
      661 aaaggacgaa acaccggaag gagttccagg tgttactgct cgagcagtaa cacctggaac
      721 tccttttttt gaattctcga cctcgagaca aatggcagta ttcatccaca attttaaaag
      781 aaaagggggg attggggggt acagtgcagg ggaaagaata gtagacataa tagcaacaga
      841 catacaaact aaagaattac aaaaacaaat tacaaaaatt caaaattttc gggtttatta
      901 cagggacagc agagatccac tttggccgcg gctcgagggg gttggggttg cgccttttcc
      961 aaggcagccc tgggtttgcg cagggacgcg gctgctctgg gcgtggttcc gggaaacgca
     1021 gcggcgccga ccctgggtct cgcacattct tcacgtccgt tcgcagcgtc acccggatct
     1081 tcgccgctac ccttgtgggc cccccggcga cgcttcctgc tccgccccta agtcgggaag
     1141 gttccttgcg gttcgcggcg tgccggacgt gacaaacgga agccgcacgt ctcactagta
     1201 ccctcgcaga cggacagcgc cagggagcaa tggcagcgcg ccgaccgcga tgggctgtgg
     1261 ccaatagcgg ctgctcagca gggcgcgccg agagcagcgg ccgggaaggg gcggtgcggg
     1321 aggcggggtg tggggcggta gtgtgggccc tgttcctgcc cgcgcggtgt tccgcattct
     1381 gcaagcctcc ggagcgcacg tcggcagtcg gctccctcgt tgaccgaatc accgacctct
     1441 ctccccaggg ggatccaccg gagcttacca tgaccgagta caagcccacg gtgcgcctcg
     1501 ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc gccgactacc
     1561 ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc gagctgcaag
     1621 aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg
     1681 ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga
     1741 tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag
     1801 gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct
     1861 cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg
     1921 ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct
     1981 acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct
     2041 ggtgcatgac ccgcaagccc ggtgcctgac gcccgcccca cgacccgcag cgcccgaccg
     2101 aaaggagcgc acgaccccat gcatcggtac ctttaagacc aatgacttac aaggcagctg
     2161 tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt cactcccaac
     2221 gaagacaaga tctgcttttt gcttgtactg ggtctctctg gttagaccag atctgagcct
     2281 gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag
     2341 tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac
     2401 ccttttagtc agtgtggaaa atctctagca gtagtagttc atgtcatctt attattcagt
     2461 atttataact tgcaaagaaa tgaatatcag agagtgagag gaacttgttt attgcagctt
     2521 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     2581 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggctctagc
     2641 tatcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     2701 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga
     2761 gctattccag aagtagtgag gaggcttttt tggaggccta gggacgtacc caattcgccc
     2821 tatagtgagt cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa
     2881 aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt
     2941 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa
     3001 tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     3061 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     3121 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     3181 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     3241 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     3301 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     3361 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     3421 tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa
     3481 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     3541 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     3601 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     3661 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     3721 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     3781 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     3841 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     3901 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     3961 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     4021 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     4081 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     4141 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     4201 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     4261 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     4321 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     4381 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     4441 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     4501 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     4561 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     4621 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     4681 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     4741 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     4801 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     4861 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     4921 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     4981 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     5041 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     5101 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     5161 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     5221 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     5281 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     5341 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa
     5401 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
     5461 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt
     5521 aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg
     5581 gataacaatt tcacacagga aacagctatg accatgatta cgccaagcgc gcaattaacc
     5641 ctcactaaag ggaacaaaag ctggagctgc aagcttaatg tagtcttatg caatactctt
     5701 gtagtcttgc aacatggtaa cgatgagtta gcaacatgcc ttacaaggag agaaaaagca
     5761 ccgtgcatgc cgattggtgg aagtaaggtg gtacgatcgt gccttattag gaaggcaaca
     5821 gacgggtctg acatggattg gacgaaccac tgaattgccg cattgcagag atattgtatt
     5881 taagtgccta gctcgataca taaacgggtc tctctggtta gaccagatct gagcctggga
     5941 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct
     6001 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt
     6061 ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg
     6121 aaaccagagg agctctctcg acgcaggact cggcttgctg aagcgcgcac ggcaagaggc
     6181 gaggggcggc gactggtgag tacgccaaaa attttgacta gcggaggcta gaaggagaga
     6241 gatgggtgcg agagcgtcag tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt
     6301 cggttaaggc cagggggaaa gaaaaaatat aaattaaaac atatagtatg ggcaagcagg
     6361 gagctagaac gattcgcagt taatcctggc ctgttagaaa catcagaagg ctgtagacaa
     6421 atactgggac agctacaacc atcccttcag acaggatcag aagaacttag atcattatat
     6481 aatacagtag caaccctcta ttgtgtgcat caaaggatag agataaaaga caccaaggaa
     6541 gctttagaca agatagagga agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc
     6601 gctgatcttc agacctggag gaggagatat gagggacaat tggagaagtg aattatataa
     6661 atataaagta gtaaaaattg aaccattagg agtagcaccc accaaggcaa agagaagagt
     6721 ggtgcagaga gaaaaaagag cagtgggaat aggagctttg ttccttgggt tcttgggagc
     6781 agcaggaagc actatgggcg cagcgtcaat gacgctgacg gtacaggcca gacaattatt
     6841 gtctggtata gtgcagcagc agaacaattt gctgagggct attgaggcgc aacagcatct
     6901 gttgcaactc acagtctggg gcatcaagca gctccaggca agaatcctgg ctgtggaaag
     6961 atacctaaag gatcaacagc tcctggggat ttggggttgc tctggaaaac tcatttgcac
     7021 cactgctgtg ccttggaatg ctagttggag taataaatct ctggaacaga tttggaatca
     7081 cacg
//
