LOCUS       sequence-170408-        7948 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7948)
  AUTHORS   Landthaler M, Gaidatzis D, Rothballer A, Chen PY, Soll SJ, Dinic L, 
            Ojo T, Hafner M, Zavolan M, Tuschl T
  TITLE     Molecular characterization of human Argonaute-containing 
            ribonucleoprotein complexes and their bound target mRNAs.
  JOURNAL   RNA. 2008 Dec;14(12):2580-96. doi: 10.1261/rna.1351608. Epub 2008 
            Oct 31.
  PUBMED    18978028
REFERENCE   2  (bases 1 to 7948)
  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.1351608"; journalName: "RNA"; date: "2008-12"; volume: 
            "14"; issue: "12"; pages: "2580-96"
COMMENT     Sequence Label: pFRT/FLAG/HA-DEST EIF2C2
FEATURES             Location/Qualifiers
     source          1..7948
                     /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"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      946..955
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             955..978
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             979..1005
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    1021..1045
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      3067..3076
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(3744..3768)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3821..3862
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3900..3917)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3906..4130
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4263..4282)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4414..4461
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             4469..5491
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4469..5491
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    5621..5742
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5658..5677)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5712..5731
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5791..5807)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5791..5807)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5804..5826)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5815..5831
                     /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(5839..5845,5846..5863,5864..5869))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5884..5905
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6022..6039)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6193..6781)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6273..6292)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6952..7743,7744..7812))
                     /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     7575..7594
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7813..7917)
                     /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 agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gagctcggat cgatccatcg atagtgtggt ggaattctga tcgccgccgc catggactac
      961 aaggacgacg atgacaagta cccttatgac gtgcccgatt acgctttgaa ccagatatca
     1021 acaagtttgt acaaaaaagc aggctccacc atgggaacca attcagtcga catgtactcg
     1081 ggagccggct cccgagtaca tgtcgacgcg tgggccggcc cacgcgtcga ctggatccgg
     1141 atgtactcgg gagccggccc cgcacttgca cctcctgcgc cgccgccccc catccaagga
     1201 tatgccttca agcctccacc tagacccgac tttgggacct ccgggagaac aatcaaatta
     1261 caggccaatt tcttcgaaat ggacatcccc aaaattgaca tctatcatta tgaattggat
     1321 atcaagccag agaagtgccc gaggagagtt aacagggaaa tcgtggaaca catggtccag
     1381 cactttaaaa cacagatctt tggggatcgg aagcccgtgt ttgacggcag gaagaatcta
     1441 tacacagcca tgccccttcc gattgggagg gacaaggtgg agctggaggt cacgctgcca
     1501 ggagaaggca aggatcgcat cttcaaggtg tccatcaagt gggtgtcctg cgtgagcttg
     1561 caggcgttac acgatgcact ttcagggcgg ctgcccagcg tcccttttga gacgatccag
     1621 gccctggacg tggtcatgag gcacttgcca tccatgaggt acacccccgt gggccgctcc
     1681 ttcttcaccg cgtccgaagg ctgctctaac cctcttggcg ggggccgaga agtgtggttt
     1741 ggcttccatc agtccgtccg gccttctctc tggaaaatga tgctgaatat tgatgtgtca
     1801 gcaacagcgt tttacaaggc acagccagta atcgagtttg tttgtgaagt tttggatttt
     1861 aaaagtattg aagaacaaca aaaacctctg acagattccc aaagggtaaa gtttaccaaa
     1921 gaaattaaag gtctaaaggt ggagataacg cactgtgggc agatgaagag gaagtaccgt
     1981 gtctgcaatg tgacccggcg gcccgccagt caccaaacat tcccgctgca gcaggagagc
     2041 gggcagacgg tggagtgcac ggtggcccag tatttcaagg acaggcacaa gttggttctg
     2101 cgctaccccc acctcccatg tttacaagtc ggacaggagc agaaacacac ctaccttccc
     2161 ctggaggtct gtaacattgt ggcaggacaa agatgtatta aaaaattaac ggacaatcag
     2221 acctcaacca tgatcagagc aactgctagg tcggcgcccg atcggcaaga agagattagc
     2281 aaattgatgc gaagtgcaag tttcaacaca gatccatacg tccgtgaatt tggaatcatg
     2341 gtcaaagatg agatgacaga cgtgactggg cgggtgctgc agccgccctc catcctctac
     2401 gggggcagga ataaagctat tgcgacccct gtccagggcg tctgggacat gcggaacaag
     2461 cagttccaca cgggcatcga gatcaaggtg tgggccattg cgtgcttcgc cccccagcgc
     2521 cagtgcacgg aagtccatct gaagtccttc acagagcagc tcagaaagat ctcgagagac
     2581 gctggcatgc ccatccaggg ccagccgtgc ttctgcaaat acgcgcaggg ggcggacagc
     2641 gtggagccca tgttccggca cctgaagaac acgtatgcgg gcctgcagct ggtggtggtc
     2701 atcctgcccg gcaagacgcc cgtgtacgcc gaggtcaagc gcgtgggaga cacggtgctg
     2761 gggatggcca cgcagtgcgt gcagatgaag aacgtgcaga ggaccacgcc acagaccctg
     2821 tccaaccttt gcctgaagat caacgtcaag ctgggaggcg tgaacaacat cctgctgccc
     2881 cagggcaggc cgccggtgtt ccagcagccc gtcatctttc tgggagcaga cgtcactcac
     2941 ccccccgccg gggatgggaa gaagccctcc attgccgccg tggtgggcag catggacgcc
     3001 caccccaatc gctactgcgc caccgtgcgc gtgcagcagc accggcagga gatcatacaa
     3061 gacctggccg ccatggtccg cgagctcctc atccagttct acaagtccac gcgcttcaag
     3121 cccacccgca tcatcttcta ccgcgacggt gtctctgaag gccagttcca gcaggttctc
     3181 caccacgagt tgctggccat ccgtgaggcc tgtatcaagc tagaaaaaga ctaccagccc
     3241 gggatcacct tcatcgtggt gcagaagagg caccacaccc ggctcttctg cactgacaag
     3301 aacgagcggg ttgggaaaag tggaaacatt ccagcaggca cgactgtgga cacgaaaatc
     3361 acccacccca ccgagttcga cttctacctg tgtagtcacg ctggcatcca ggggacaagc
     3421 aggccttcgc actatcacgt cctctgggac gacaatcgtt tctcctctga tgagctgcag
     3481 atcctaacct accagctgtg tcacacctac gtgcgctgca cacgctccgt gtccatccca
     3541 gcgccagcat actacgctca cctggtggcc ttccgggcca ggtaccacct ggtggataag
     3601 gaacatgaca gtgctgaagg aagccatacc tctgggcaga gtaacgggcg agaccaccaa
     3661 gcactggcca aggcggtcca ggttcaccaa gacactctgc gcaccatgta ctttgcttga
     3721 gcggccgcac tcgagatatc tagacccagc tttcttgtac aaagtggttg atatccagca
     3781 cagtggcggc cgctcaagtc tagagggccc gcggttcgaa ggtaagccta tccctaaccc
     3841 tctcctcggt ctcgattcta cgctcgagtc tagagggccc gtttaaaccc gctgatcagc
     3901 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     3961 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     4021 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     4081 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
     4141 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     4201 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     4261 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     4321 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     4381 aaaacttgat tagggtgatg gttcacgtac ctagaagttc ctattccgaa gttcctattc
     4441 tctagaaagt ataggaactt ccttggccaa aaagcctgaa ctcaccgcga cgtctgtcga
     4501 gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg atgcagctct cggagggcga
     4561 agaatctcgt gctttcagct tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag
     4621 ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg cactttgcat cggccgcgct
     4681 cccgattccg gaagtgcttg acattgggga attcagcgag agcctgacct attgcatctc
     4741 ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa accgaactgc ccgctgttct
     4801 gcagccggtc gcggaggcca tggatgcgat cgctgcggcc gatcttagcc agacgagcgg
     4861 gttcggccca ttcggaccgc aaggaatcgg tcaatacact acatggcgtg atttcatatg
     4921 cgcgattgct gatccccatg tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc
     4981 gtccgtcgcg caggctctcg atgagctgat gctttgggcc gaggactgcc ccgaagtccg
     5041 gcacctcgtg cacgcggatt tcggctccaa caatgtcctg acggacaatg gccgcataac
     5101 agcggtcatt gactggagcg aggcgatgtt cggggattcc caatacgagg tcgccaacat
     5161 cttcttctgg aggccgtggt tggcttgtat ggagcagcag acgcgctact tcgagcggag
     5221 gcatccggag cttgcaggat cgccgcggct ccgggcgtat atgctccgca ttggtcttga
     5281 ccaactctat cagagcttgg ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg
     5341 atgcgacgca atcgtccgat ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag
     5401 aagcgcggcc gtctggaccg atggctgtgt agaagtactc gccgatagtg gaaaccgacg
     5461 ccccagcact cgtccgaggg caaaggaata gcacgtacta cgagatttcg attccaccgc
     5521 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     5581 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     5641 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     5701 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     5761 gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     5821 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     5881 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     5941 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     6001 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     6061 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     6121 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     6181 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     6241 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     6301 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     6361 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     6421 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     6481 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     6541 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     6601 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     6661 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     6721 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     6781 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     6841 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     6901 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     6961 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     7021 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     7081 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     7141 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     7201 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     7261 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     7321 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     7381 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     7441 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     7501 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     7561 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     7621 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     7681 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     7741 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     7801 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     7861 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     7921 atttccccga aaagtgccac ctgacgtc
//
