LOCUS       sequence-75895-e        5659 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FLAG, His tagged BmAgo3 in silkworm cell line BmN4 or 
            insect cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5659)
  AUTHORS   Kawaoka S, Hayashi N, Suzuki Y, Abe H, Sugano S, Tomari Y, Shimada 
            T, Katsuma S
  TITLE     The Bombyx ovary-derived cell line endogenously expresses 
            PIWI/PIWI-interacting RNA complexes.
  JOURNAL   RNA. 2009 Jul;15(7):1258-64. doi: 10.1261/rna.1452209. Epub 2009 May
            21.
  PUBMED    19460866
REFERENCE   2  (bases 1 to 5659)
  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: 
            "10.1261/rna.1452209"; journalName: "RNA"; date: "2009-07"; volume: 
            "15"; issue: "7"; pages: "1258-64"
COMMENT     Sequence Label: pIZ-Flag6His-BmAgo3
FEATURES             Location/Qualifiers
     source          1..5659
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        5..552
                     /gene="immediate-early gene 2 from the Orgyia pseudotsugata
                     multicapsid nucleopolyhedrovirus"
                     /label=OpIE-2 promoter
                     /note="strong constitutive baculovirus promoter for insect 
                     cell expression"
     CDS             588..611
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             612..629
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3446..3487
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             3497..3514
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    3532..3661
                     /gene="immediate-early gene 2 from the Orgyia pseudotsugata
                     multicapsid nucleopolyhedrovirus"
                     /label=OpIE-2 poly(A) signal
                     /note="baculovirus polyadenylation signal"
     rep_origin      complement(3789..4377)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3869..3888)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        4449..4996
                     /gene="immediate-early gene 2 from the Orgyia pseudotsugata
                     multicapsid nucleopolyhedrovirus"
                     /label=OpIE-2 promoter
                     /note="strong constitutive baculovirus promoter for insect 
                     cell expression"
     promoter        5025..5072
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        5064..5082
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      5086..5095
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5092..5466
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    join(5530..5659,1..5)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5567..5586)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5621..5640
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 ggatcatgat gataaacaat gtatggtgct aatgttgctt caacaacaat tctgttgaac
       61 tgtgttttca tgtttgccaa caagcacctt tatactcggt ggcctcccca ccaccaactt
      121 ttttgcactg caaaaaaaca cgcttttgca cgcgggccca tacatagtac aaactctacg
      181 tttcgtagac tattttacat aaatagtcta caccgttgta tacgctccaa atacactacc
      241 acacattgaa cctttttgca gtgcaaaaaa gtacgtgtcg gcagtcacgt aggccggcct
      301 tatcgggtcg cgtcctgtca cgtacgaatc acattatcgg accggacgag tgttgtctta
      361 tcgtgacagg acgccagctt cctgtgttgc taaccgcagc cggacgcaac tccttatcgg
      421 aacaggacgc gcctccatat cagccgcgcg ttatctcatg cgcgtgaccg gacacgaggc
      481 gcccgtcccg cttatcgcgc ctataaatac agcccgcaac gatctggtaa acacagttga
      541 acagcatctg ttcgaattta aagcttggta ccgagctcgg atccatggat tacaaggatg
      601 acgacgataa gcatcatcat catcatcata tggcagaccc aggcaaaggc cgggggcgaa
      661 gcttagccct tttacaggca ctgaagaagt cacaaatgat ggactctccc tcacaatctg
      721 agagtcagtc tcctgaaagt acaccagaac agagtacagc tccaagtacc attgcaagtg
      781 caactcctag tacttcagga gtgagtatag gcggtagagg acgcgcagca gcattgatgc
      841 tggctaaaat gcagcagaaa ccaggctcca caactccagc aatttttgtg ccaccttcat
      901 caacttctgc accaacagct ggaactggac gtggattcaa attgcttcag aacttgcaag
      961 ccagtcaaaa agcaagcagt caaatagcaa gcagtcaagt aacgagcagt gcacaaagcg
     1021 atattaaaga tttgactgaa aaaatgtctg aaacttcagt ttcagcacag gcatcttcag
     1081 ttgccaagaa caagtatttc agggaggtga aagacactcc tccagtcgta aagaaaggtg
     1141 aaacgggtgt gcctatagaa gtgacctgta actacattta tctcaatttc aaagagaaca
     1201 ttgtttttga gtatgaagtg aagtttgaac cagatcaaga ctataagcat ttgcgattca
     1261 aattgctcaa tgagcacatt gaatacttta aggagaaaac attcgacggt accaccctct
     1321 acgtgccaca cgagctgccc gatgcagtcc gtaatttagt atctactaac ccatatgacc
     1381 aaagcaaagt aaacgtttct attatatttc gtcgcacacg tcgtttgagc gaaatgattc
     1441 acatatacaa tgtgatgttc aaatgtataa tgaaggattt gaaattgatt cgcttcggac
     1501 gccaacatta caatgagcac gcagccatac agatacctca gcataaatta gaagtttggc
     1561 ctggatacgt aacagcagta gacgaatacg aaggcggtct gatgttaaca ttagattcga
     1621 ctcatcgtgt attacgaacg caaaccgttc tctctctgat caaagaggtt gtgcaaactg
     1681 aaggagctaa ttggaaacgt aagatgacag atatactaat aggagcctct gttatgacta
     1741 cttataacaa gaaattgttt agagttgata caattgacga taaaatgagt ccgagatcaa
     1801 catttgagaa aacagagaaa ggcgaaacta tacagattag ttttatagat tattataaga
     1861 agaattatgg tatcgagata atggactggg atcaaccctt actcatttca agagacacaa
     1921 aacgtatgcc cggatcagac actccaacgg atttcatgat atgcctcatt ccggagttat
     1981 gtcaattgac tggtttaaca gatgatcaac gcagtaactt tagattaatg aaagatgtag
     2041 ctacttacac gaggatcact ccaaatcagc gccatgccgc ttttaagaag tacatagaaa
     2101 gtgtaatgaa aaacgaaact gccaagagtc gtctagcggg ttggggtctc agtatagcac
     2161 cggaaactgt gaacttaaca gctcgtaccc tgccaccaga gacactgtat tttggagaca
     2221 atgttagagt tcccggcaaa ccgaatgcag agtggaactc tgaggtgact aagcacagtg
     2281 tgatgcaagc tgtggatatc atgagatggg ttctgttgtt cactcagagg gataagcaag
     2341 ttgctatgga ctttcttagt acactcaaac gaaattgtcg accaatgggt attatggttt
     2401 ctgacgctga attggtaccc ctcgccaatg atagaacaga tacgtacgta ctggcattga
     2461 agaagtgcat aacgtcgtct gtacaattag tcgttgcgat atgttctacg aagagagatg
     2521 acagatacgc ggccattaag aaagtttgtt gtgctgacaa ccccgtgccc tcacaggtta
     2581 taaatgcacg tacgttaatg aacacgaaca agatcagatc gatcactcag aagattttgc
     2641 ttcagttgaa ttgcaaactg ggcggtacgc tttggagcat cagcataccg tttaagagcg
     2701 caatgattgt aggcatcgat tcgtatcacg acccatccag gaggaaccgg agcgtttgca
     2761 gttttgtagc atcctataat caaagcatga cgctttggta ttcgaaagtg atcttccaag
     2821 agaagggtca agaaatagtt gatggcttga aatgttgtct cgttgatgcg ttaactcatt
     2881 atttgagatc aaacggacag cttccggaca ggatcatcat atacagagac ggtgtaggcg
     2941 acgggcagct gaagttattg cagcaatacg agatacctca aatgaagatt tgcttcacga
     3001 tattgggatc gaattaccag cctaccctga cctatgttgt cgtacaaaaa cgtatcaaca
     3061 ctaggatctt cctgaagtcg agagacggct acgacaatcc gaaccccggg accgtggtcg
     3121 atcactgtat aacgagacgg gactggtacg atttcctgat cgtctcgcag aaagtgaccc
     3181 agggcactgt caccccgacc cattacgtgg tggtctacga cgacagcggg atcaccccgg
     3241 accagtgtca gcgtttgacg tacaagatgt gtcacttgta ctacaattgg ccggggacgg
     3301 tgcgcgtgcc ggcgccatgt cagtatgcgc acaaactctc gtatctagtg ggacagtgcg
     3361 tgcacgcgca gccgtctgac gttttagtcg ataagctgtt ctttttgtag gcggccgctc
     3421 gagtctagag ggcccgcggt tcgaaggtaa gcctatccct aaccctctcc tcggtctcga
     3481 ttctacgcgt accggtcatc atcaccatca ccattgagtt tatctgacta aatcttagtt
     3541 tgtattgtca tgttttaata caatatgtta tgtttaaata tgtttttaat aaattttata
     3601 aaataatttc aacttttatt gtaacaacat tgtccattta cacactcctt tcaagcgcgt
     3661 gggatcgatg ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     3721 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     3781 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     3841 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     3901 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     3961 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     4021 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     4081 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     4141 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     4201 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     4261 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     4321 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     4381 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     4441 attttggtca tgatgataaa caatgtatgg tgctaatgtt gcttcaacaa caattctgtt
     4501 gaactgtgtt ttcatgtttg ccaacaagca cctttatact cggtggcctc cccaccacca
     4561 acttttttgc actgcaaaaa aacacgcttt tgcacgcggg cccatacata gtacaaactc
     4621 tacgtttcgt agactatttt acataaatag tctacaccgt tgtatacgct ccaaatacac
     4681 taccacacat tgaacctttt tgcagtgcaa aaaagtacgt gtcggcagtc acgtaggccg
     4741 gccttatcgg gtcgcgtcct gtcacgtacg aatcacatta tcggaccgga cgagtgttgt
     4801 cttatcgtga caggacgcca gcttcctgtg ttgctaaccg cagccggacg caactcctta
     4861 tcggaacagg acgcgcctcc atatcagccg cgcgttatct catgcgcgtg accggacacg
     4921 aggcgcccgt cccgcttatc gcgcctataa atacagcccg caacgatctg gtaaacacag
     4981 ttgaacagca tctgttcgaa ttaattcgga tctctgcagc acgtgttgac aattaatcat
     5041 cggcatagta tatcggcata gtataatacg actcactata ggagggccac catggccaag
     5101 ttgaccagtg ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg
     5161 accgaccggc tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg
     5221 gacgacgtga ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg
     5281 gcctgggtgt gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc
     5341 acgaacttcc gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg
     5401 cgggagttcg ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag
     5461 gactgaccga cgccgaccaa caccgccggt ccgacggcgg cccacgggtc ccaggggggt
     5521 cgacctcgaa acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     5581 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     5641 aatgtatctt atcatgtct
//
