LOCUS       sequence-290287-        7444 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Insect Expression of DmMe31B-Q294AAN296F-dsRNAres.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7444)
  AUTHORS   Chen Y, Boland A, Kuzuoglu-Ozturk D, Bawankar P, Loh B, Chang CT, 
            Weichenrieder O, Izaurralde E
  TITLE     A DDX6-CNOT1 complex and W-binding pockets in CNOT9 reveal direct 
            links between miRNA target recognition and silencing.
  JOURNAL   Mol Cell. 2014 Jun 5;54(5):737-50. doi: 
            10.1016/j.molcel.2014.03.034. Epub 2014 Apr 24.
  PUBMED    24768540
REFERENCE   2  (bases 1 to 7444)
  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.2014.03.034"; journalName: "Mol Cell"; date: 
            "2014-06-5- 5"; volume: "54"; issue: "5"; pages: "737-50"
COMMENT     Sequence Label: pAc5.1B-EGFP-DmMe31B-Q294AN296F-dsRNAres_W
FEATURES             Location/Qualifiers
     source          1..7444
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        56..2568
                     /label=Ac5 promoter
                     /note="Drosophila melanogaster actin 5C promoter"
     CDS             join(2588..2590,2591..2593,2594..3304)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2588..2590,2591..2593,2594..3304)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2633..2654)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2894..2913)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3241..3262
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             4729..4770
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             4780..4797
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4820..4837)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4870..5004
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4894..4913)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4948..4967
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(5626..6214)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5706..5725)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6385..7176,7177..7245))
                     /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     7008..7027
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7246..7350)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7418..7436
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 cttcaaggaa ttaattctat attctaaaaa cacaaatgat acttctaaaa aaaatcatga
       61 atggcatcaa ctctgaatca aatctttgca gatgcaccta cttctcattt ccactgtcac
      121 atcatttttc cagatctcgc tgcctgttat gtggcccaca aaccaagaca cgttttatgg
      181 ccattaaagc tggctgatcg tcgccaaaca ccaaatacat atcaatatgt acattcgaga
      241 aagaagcgat caaagaagcg tcttcgggcg agtaggagaa tgcggaggag aaggagaacg
      301 agctgatcta gtatctctcc acaatccaat gccaactgac caactggcca tattcggagc
      361 aatttgaagc caatttccat cgcctggcga tcgctccatt cttggctata tgtttttcac
      421 cgttcccggg gccattttca aagactcgtc ggtaagataa gattgtgtca ctcgctgtct
      481 ctcttcattt gtcgaagaat gctgaggaat ttcgcgatga cgtcggcgag tattttgaag
      541 aatgagaata atttgtattt atacgaaaat cagttagtgg aattttctac aaaaacatgt
      601 tatctataga taattttgtt gcaaaatatg ttgactatga caaagattgt atgtatatac
      661 ctttaatgta ttctcatttt cttatgtatt tataatggca atgatgatac tgatgatatt
      721 ttaagatgat gccagaccac aggctgattt ctgcgtcttt tgccgaacgc agtgcatgtg
      781 cggttgttgt tttttggaat agtttcaatt ttcggactgt ccgctttgat ttcagtttct
      841 tggcttattc aaaaagcaaa gtaaagccaa aaaagcgaga tggcaatacc aaatgcggca
      901 aaacggtagt ggaaggaaag gggtgcgggg cagcggaagg aagggtgggg cggggcgtgg
      961 cggggtctgt ggctgggcgc gacgtcaccg acgttggagc cactcctttg accatgtgtg
     1021 cgtgtgtgta ttattcgtgt ctcgccactc gccggttgtt tttttctttt tatctcgctc
     1081 tctctagcgc catctcgtac gcatgctcaa cgcaccgcat gttgccgtgt cctttatgcg
     1141 tcattttggc tcgaaatagg caattattta aacaaagatt agtcaacgaa aacgctaaaa
     1201 taaataagtc tacaatatgg ttacttattg ccatgtgtgt gcagccaacg atagcaacaa
     1261 aagcaacaac acagtggctt tccctctttc actttttgtt tgcaagcgcg tgcgagcaag
     1321 acggcacgac cggcaaacgc aattacgctg acaaagagca gacgaagttt tggccgaaaa
     1381 acatcaaggc gcctgatacg aatgcatttg caataacaat tgcgatattt aatattgttt
     1441 atgaagctgt ttgacttcaa aacacacaaa aaaaaaaata aaacaaatta tttgaaagag
     1501 aattaggaat cggacagctt atcgttacgg gctaacagca caccgagacg aaatagctta
     1561 cctgacgtca cagcctctgg aagaactgcc gccaagcaga gagagagaga aaaagaggga
     1621 gagcagctta gaccgcatgt gcttgtgtgt gaggcgtctc tctcttcgtc tcctgtttgc
     1681 gcaaacgcat agactgcact gagaaaatcg attacctatt ttttatgaat gaatatttgc
     1741 actattacta ttcaaaacta ttaagatagc aatcacattc aatagccaaa tactatacca
     1801 cctgagcgat gcaacgaaat gatcaatttg agcaaaaatg ctgcatattt aggacggcat
     1861 cattatagaa atgcttcttg ctgtgtactt ttctctcgtc tggcagctgt ttcgccgtta
     1921 ttgttaaaac cggcttaagt taggtgtgtt ttctacgact agtgatgccc ctactagaag
     1981 atgtgtgttg cacaaatgtc cctgaataac caatttgaag tgcagatagc agtaaacgta
     2041 agctaatatg aatattattt aactgtaatg ttttaatatc gctggacatt actaataaac
     2101 ccactataaa cacatgtaca tatgtatgtt ttggcataca atgagtagtt ggggaaaaaa
     2161 tgtgtaaaag caccgtgacc atcacagcat aaagataacc agctgaagta tcgaatatga
     2221 gtaaccccca aattgaatca catgccgcaa ctgataggac ccatggaagt acactcttca
     2281 tggcgatata caagacacac acaagcacga acacccagtt gcggaggaaa ttctccgtaa
     2341 atgaaaaccc aatcggcgaa caattcatac ccatatatgg taaaagtttt gaacgcgact
     2401 tgagagcgga gagcattgcg gctgataagg ttttagcgct aagcgggctt tataaaacgg
     2461 gctgcgggac cagttttcat atcactaccg tttgagttct tgtgctgtgt ggatactcct
     2521 cccgacacaa agccgctcca tcagccagca gtcgtctaat ccagagaccc cggatcgggg
     2581 taccaacatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     2641 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     2701 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     2761 gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca
     2821 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     2881 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     2941 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     3001 ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca
     3061 gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     3121 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     3181 caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     3241 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     3301 caagccaagc ttgatgatga ccgagaaact gaactccggc cataccaacc tgacttctaa
     3361 aggaataatt aacgacctgc agatagccgg taatacttca gatgacatgg ggtggaagtc
     3421 taaattaaaa ttaccaccga aagataatcg gtttaagacg acagacgtta cagacacacg
     3481 gggaaacgag tttgaagaat tctgtttaaa gcgggagttg ctgatgggaa tctttgaaaa
     3541 aggttgggaa cggccatcac cgatacaaga agctgcaata ccaattgctc tgtcgggtaa
     3601 ggacgtccta gccagagcaa aaaatggaac gggaaaaacc ggcgcgtatt gtatacctgt
     3661 cctggagcag atagatccta caaaagatta tatacaagcc ttggttatgg ttccaacacg
     3721 ggaactggcc ttgcagacat cccaaatttg catagaatta gccaagcacc tggacattag
     3781 agttatggtt acgacaggtg gaacaatctt gaaagacgat atattgcgaa tataccagaa
     3841 ggtccagcta atcatcgcaa caccaggccg gattttggac cttatggaca aaaaggtagc
     3901 agacatgagt cactgtcgaa ttctggtatt agacgaagca gacaaattgc tatcactaga
     3961 cttccaggga atgttagacc acgtcatctt aaaactaccc aaagaccctc aaatactatt
     4021 gttctcggcc accttcccgt taactgtaaa aaatttcatg gagaagcatt tacgcgagcc
     4081 ttacgagatc aacctgatgg aggagctaac actgaagggt gtcactcagt actatgcatt
     4141 cgtacaggag cgccagaaag tacactgctt gaatacgctc ttttcgaagc tggcgatcgc
     4201 tcagtctatc atattctgca attccacgca acgtgtcgag ctattggcca aaaagatcac
     4261 agagcttgga tattgctgtt actatataca cgctaagatg gcccaggcac acagaaatcg
     4321 agtattccac gatttccgcc aaggactctg tcgcaatctg gtgtgctccg acctgttcac
     4381 ccgtggtatc gacgtgcagg ccgtgaatgt agtgatcaat ttcgatttcc cacgaatggc
     4441 agagacatac ttgcatcgaa ttggtcgctc aggacgtttc ggtcatttgg gtattgctat
     4501 taatctgata acctacgagg atcggtttga tctgcatcgg attgaaaaag aacttggaac
     4561 cgaaataaag cctattccga aggtcataga ccccgccctt tatgtcgcaa acgttggcgc
     4621 atccgttggc gatacttgca acaacagtga tctgaataat tctgcgaacg aggagggcaa
     4681 cgttagcaaa taagcggccg ctcgagtcta gagggcccgc ggttcgaagg taagcctatc
     4741 cctaaccctc tcctcggtct cgattctacg cgtaccggtc atcatcacca tcaccattga
     4801 gtttaaaccc gctgatcagc ctcgactgtg ccttctaagg cctgagctcg ctgatcagcc
     4861 tcgatcgagg atccagacat gataagatac attgatgagt ttggacaaac cacaactaga
     4921 atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc
     4981 attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt
     5041 cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtatggct
     5101 gattatgatc agtcgaccga tgcccttgag agccttcaac ccagtcagct ccttccggtg
     5161 ggcgcggggc atgactatcg tcgccgcact tatgactgtc ttctttatca tgcaactcgt
     5221 aggacaggtg ccggcagcgc tctgggtcat tttcggcgag gaccgctttc gctggagcgc
     5281 gacgatgatc ggcctgtcgc ttgcggtatt cggaatcttg cacgccctcg ctcaagcctt
     5341 cgtcactggt cccgccacca aacgtttcgg cgagaagcag gccattatcg ccggcatggc
     5401 ggccgacgcg ctgggctacg tcttgctggc gttcgcgacg cgaggctgga tggccttccc
     5461 cattatgatt cttctcgctt ccggcggcat cgggatgccc gcgttgcagg ccatgctgtc
     5521 caggcaggta gatgacgacc atcagggaca gcttcaagga tcgctcgcgg ctcttaccag
     5581 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     5641 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     5701 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     5761 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     5821 atgctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     5881 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     5941 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     6001 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     6061 tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     6121 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     6181 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     6241 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     6301 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     6361 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     6421 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     6481 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     6541 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     6601 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     6661 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct gcaggcatcg tggtgtcacg
     6721 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     6781 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     6841 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     6901 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     6961 atagtgtatg cggcgaccga gttgctcttg cccggcgtca acacgggata ataccgcgcc
     7021 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     7081 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     7141 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     7201 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     7261 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     7321 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     7381 ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt
     7441 tcgt
//
