LOCUS       sequence-290872-        6621 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of TISU-HA-hRluc-V5-6xbsMS2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6621)
  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 6621)
  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: pcDNA3.1-TISU-HA-hRLuc-V5-6xbsMS2_AC
FEATURES             Location/Qualifiers
     source          1..6621
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             844..870
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             871..1800
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="ASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAENA
                     VIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAWF
                     ELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDIA
                     LIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPREI
                     PLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKVK
                     GLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     CDS             1801..1842
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     misc_RNA        1942..1960
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        1981..1999
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        2020..2038
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        2059..2077
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        2098..2116
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        2137..2155
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     promoter        complement(2177..2195)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2178..2195)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2215..2232)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2221..2445
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2491..2919
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2578..2597)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2788..2809
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2928..2948)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2933..3262
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3113..3248
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3175..3194
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3329..4123
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3383..3402)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3993..4012
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4297..4418
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4334..4353)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4388..4407
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4467..4483)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4467..4483)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4480..4502)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4491..4507
                     /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)."
     promoter        complement(join(4515..4521,4522..4539,4540..4545))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4560..4581
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4698..4715)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4869..5454)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4949..4968)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5625..6416,6417..6485))
                     /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     6248..6267
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6486..6590)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact caagatggcg
      841 gcatacccat atgatgttcc agattacgct gcttccaagg tgtacgaccc cgagcaacgc
      901 aaacgcatga tcactgggcc tcagtggtgg gctcgctgca agcaaatgaa cgtgctggac
      961 tccttcatca actactatga ttccgagaag cacgccgaga acgccgtgat ttttctgcat
     1021 ggtaacgctg cctccagcta cctgtggagg cacgtcgtgc ctcacatcga gcccgtggct
     1081 agatgcatca tccctgatct gatcggaatg ggtaagtccg gcaagagcgg gaatggctca
     1141 tatcgcctcc tggatcacta caagtacctc accgcttggt tcgagctgct gaaccttcca
     1201 aagaaaatca tctttgtggg ccacgactgg ggggcttgtc tggcctttca ctactcctac
     1261 gagcaccaag acaagatcaa ggccatcgtc catgctgaga gtgtcgtgga cgtgatcgag
     1321 tcctgggacg agtggcctga catcgaggag gatatcgccc tgatcaagag cgaagagggc
     1381 gagaaaatgg tgcttgagaa taacttcttc gtcgagacca tgctcccaag caagatcatg
     1441 cggaaactgg agcctgagga gttcgctgcc tacctggagc cattcaagga gaagggcgag
     1501 gttagacggc ctaccctctc ctggcctcgc gagatccctc tcgttaaggg aggcaagccc
     1561 gacgtcgtcc agattgtccg caactacaac gcctaccttc gggccagcga cgatctgcct
     1621 aagatgttca tcgagtccga ccctgggttc ttttccaacg ctattgtcga gggagctaag
     1681 aagttcccta acaccgagtt cgtgaaggtg aagggcctcc acttcagcca ggaggacgct
     1741 ccagatgaaa tgggtaagta catcaagagc ttcgtggagc gcgtgctgaa gaacgagcag
     1801 ggtaagccta tccctaaccc tctcctcggt ctcgattcta ccgtaagcgg ccgctcgctt
     1861 tcttgctgtc caatttctat taaaggttcc tttgttccct aagtccaact accaaactgg
     1921 gtctagctct agctgtagaa aacatgagga tcacccatgt ctgctggacg actgtagaaa
     1981 acatgaggat cacccatgtc tgctgtctag ctgtagaaaa catgaggatc acccatgtct
     2041 gctggacgac tgtagaaaac atgaggatca cccatgtctg ctgtctagct gtagaaaaca
     2101 tgaggatcac ccatgtctgc tggacgactg tagaaaacat gaggatcacc catgtctgct
     2161 gtctagaggg ccctattcta tagtgtcacc taaatgctag agctcgctga tcagcctcga
     2221 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     2281 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     2341 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     2401 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     2461 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     2521 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     2581 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     2641 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     2701 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     2761 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     2821 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     2881 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     2941 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3001 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3061 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     3121 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     3181 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     3241 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     3301 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     3361 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     3421 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     3481 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
     3541 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     3601 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     3661 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     3721 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     3781 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     3841 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     3901 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     3961 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     4021 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     4081 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     4141 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     4201 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     4261 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
     4321 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     4381 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc
     4441 tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     4501 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     4561 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     4621 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     4681 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     4741 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     4801 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     4861 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     4921 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     4981 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     5041 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     5101 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     5161 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     5221 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     5281 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     5341 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     5401 agcggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     5461 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     5521 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     5581 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     5641 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     5701 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     5761 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     5821 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     5881 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     5941 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     6001 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     6061 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     6121 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     6181 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     6241 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     6301 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     6361 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     6421 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     6481 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     6541 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     6601 cgaaaagtgc cacctgacgt c
//
