LOCUS       sequence-290856-        7085 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Insect Expression of 5BoxB.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7085)
  AUTHORS   Kuzuoglu-Ozturk D, Bhandari D, Huntzinger E, Fauser M, Helms S, 
            Izaurralde E
  TITLE     miRISC and the CCR4-NOT complex silence mRNA targets independently 
            of 43S ribosomal scanning.
  JOURNAL   EMBO J. 2016 Jun 1;35(11):1186-203. doi: 10.15252/embj.201592901. 
            Epub 2016 Mar 23.
  PUBMED    27009120
REFERENCE   2  (bases 1 to 7085)
  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: 
            "10.15252/embj.201592901"; journalName: "EMBO J"; date: "2016-06-1- 
            1"; volume: "35"; issue: "11"; pages: "1186-203"
COMMENT     Sequence Label: pAc5.1C-no5UTR-Fluc-5BoxB_AC
FEATURES             Location/Qualifiers
     source          1..7085
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             2487..4142
                     /codon_start=1
                     /gene="luc"
                     /product="firefly luciferase"
                     /label=luciferase
                     /translation="MAEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTD
                     AHIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVA
                     PANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQ
                     SMYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSH
                     ARDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDY
                     KIQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQG
                     YGLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIM
                     SGYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESI
                     LLQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGV
                     VFVDEVPKGLTGKLDARKIREILIKAKKGGKSKL"
     primer_bind     complement(2556..2574)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     protein_bind    4189..4207
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     protein_bind    4231..4249
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     protein_bind    4255..4273
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     protein_bind    4313..4331
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     protein_bind    4337..4355
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     CDS             4391..4432
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             4442..4459
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4482..4499)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4532..4666
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4556..4575)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4610..4629
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(5267..5855)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5347..5366)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6026..6817,6818..6886))
                     /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     6649..6668
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6887..6991)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7059..7077
                     /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 atcactatgg ccgaagacgc caaaaacata aagaaaggcc
     2521 cggcaccatt ctatccacta gaggatggaa ccgctggaga gcaactgcat aaggctatga
     2581 agagatacgc cctggttcct ggaacaattg cttttacaga tgcacatatc gaggtgaaca
     2641 tcacgtacgc ggaatacttc gaaatgtccg ttcggttggc agaagctatg aaacgatatg
     2701 ggctgaatac aaatcacaga atcgtcgtat gcagtgaaaa ctctcttcaa ttctttatgc
     2761 cggtgttggg cgcgttattt atcggagttg cagttgcgcc cgcgaacgac atttataatg
     2821 aacgtgaatt gctcaacagt atgaacattt cgcagcctac cgtagtgttt gtttccaaaa
     2881 aggggttgca aaaaattttg aacgtgcaaa aaaaattacc aataatccag aaaattatta
     2941 tcatggattc taaaacggat taccagggat ttcagtcgat gtacacgttc gtcacatctc
     3001 atctacctcc cggttttaat gaatacgatt ttgtaccaga gtcctttgat cgtgacaaaa
     3061 caattgcact gataatgaac tcctctggat ctactgggtt acctaagggt gtggcccttc
     3121 cgcatagaac tgcctgcgtc agattctcgc atgccagaga tcctattttt ggcaatcaaa
     3181 tcattccgga tactgcgatt ttaagtgttg ttccattcca tcacggtttt ggaatgttta
     3241 ctacactcgg atatttgata tgtggatttc gagtcgtctt aatgtataga tttgaagaag
     3301 agctgttttt acgatccctt caggattaca aaattcaaag tgcgttgcta gtaccaaccc
     3361 tattttcatt cttcgccaaa agcactctga ttgacaaata cgatttatct aatttacacg
     3421 aaattgcttc tgggggcgca cctctttcga aagaagtcgg ggaagcggtt gcaaaacgct
     3481 tccatcttcc agggatacga caaggatatg ggctcactga gactacatca gctattctga
     3541 ttacacccga gggggatgat aaaccgggcg cggtcggtaa agttgttcca ttttttgaag
     3601 cgaaggttgt ggatctggat accgggaaaa cgctgggcgt taatcagaga ggcgaattat
     3661 gtgtcagagg acctatgatt atgtccggtt atgtaaacaa tccggaagcg accaacgcct
     3721 tgattgacaa ggatggatgg ctacattctg gagacatagc ttactgggac gaagacgaac
     3781 acttcttcat agttgaccgc ttgaagtctt taattaaata caaagggtat caggtggccc
     3841 ccgctgaatt ggaatcgata ttgttacaac accccaacat cttcgacgcg ggcgtggcag
     3901 gtcttcccga cgatgacgcc ggtgaacttc ccgccgccgt tgttgttttg gagcacggaa
     3961 agacgatgac ggaaaaagag atcgtggatt acgtcgccag tcaagtaaca accgcgaaaa
     4021 agttgcgcgg aggagttgtg tttgtggacg aagtaccgaa aggtcttacc ggaaaactcg
     4081 acgcaagaaa aatcagagag atcctcataa aggccaagaa gggcggaaag tccaaattgt
     4141 aagaattcgc tagcttccct aagtccaact accaaactgg ggattcctgg gccctgaaga
     4201 agggcccctc gactagtcca aatacaaact gggccctgaa gaagggccca tatagggccc
     4261 tgaagaaggg ccctatcgag gatattatct cgactaagtc caactacaaa ctgggccctg
     4321 aagaagggcc catatagggc cctgaagaag ggccctatcg aggatattat ctcgaggtca
     4381 cccattcgaa ggtaagccta tccctaaccc tctcctcggt ctcgattcta cgcgtaccgg
     4441 tcatcatcac catcaccatt gagtttaaac ccgctgatca gcctcgactg tgccttctaa
     4501 ggcctgagct cgctgatcag cctcgatcga ggatccagac atgataagat acattgatga
     4561 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     4621 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     4681 cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa
     4741 cctctacaaa tgtggtatgg ctgattatga tcagtcgacc gatgcccttg agagccttca
     4801 acccagtcag ctccttccgg tgggcgcggg gcatgactat cgtcgccgca cttatgactg
     4861 tcttctttat catgcaactc gtaggacagg tgccggcagc gctctgggtc attttcggcg
     4921 aggaccgctt tcgctggagc gcgacgatga tcggcctgtc gcttgcggta ttcggaatct
     4981 tgcacgccct cgctcaagcc ttcgtcactg gtcccgccac caaacgtttc ggcgagaagc
     5041 aggccattat cgccggcatg gcggccgacg cgctgggcta cgtcttgctg gcgttcgcga
     5101 cgcgaggctg gatggccttc cccattatga ttcttctcgc ttccggcggc atcgggatgc
     5161 ccgcgttgca ggccatgctg tccaggcagg tagatgacga ccatcaggga cagcttcaag
     5221 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     5281 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     5341 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     5401 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     5461 aatgctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     5521 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     5581 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     5641 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     5701 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     5761 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     5821 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     5881 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     5941 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     6001 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     6061 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     6121 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     6181 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     6241 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     6301 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tgcaggcatc gtggtgtcac
     6361 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6421 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     6481 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     6541 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     6601 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aacacgggat aataccgcgc
     6661 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     6721 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     6781 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     6841 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     6901 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     6961 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     7021 tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct
     7081 ttcgt
//
