LOCUS       sequence-6802-ex        7896 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 7896)
  AUTHORS   Meister G, Landthaler M, Patkaniowska A, Dorsett Y, Teng G, Tuschl T
  TITLE     Human Argonaute2 mediates RNA cleavage targeted by miRNAs and 
            siRNAs.
  JOURNAL   Mol Cell. 2004 Jul 23. 15(2):185-97.
  PUBMED    15260970
REFERENCE   2  (bases 1 to 7896)
  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"; journalName: "Mol Cell. 
            2004 Jul 23. 15(2):185-97."
COMMENT     Sequence Label: pIRESneo-FLAG/HA Ago4
FEATURES             Location/Qualifiers
     source          1..7896
                     /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      921..930
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             930..953
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             954..980
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     intron          3652..3881
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     3864..3883
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(3939..4513,4514..4516,4517..4525)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4106..4123)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4333..4352
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4548..5351
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4611..4630)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5221..5240
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5383..5400)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5389..5613
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     5660..5679
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5739..5755)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5739..5755)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5752..5774)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5763..5779
                     /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(5787..5793,5794..5811,5812..5817))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5832..5853
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5970..5987)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6141..6729)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6221..6240)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6900..7691,7692..7760))
                     /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     7523..7542
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7761..7865)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct 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 atgggtggac 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 gcttggtacc
      901 gagctcggat cgatatcgcc gccgccatgg actacaagga cgacgatgac aagtaccctt
      961 atgacgtgcc cgattacgct agcggccgca tggaggcgct gggacccgga cctccggcta
     1021 gcctgtttca gccacctcgt cgtcctggcc ttggaactgt tggaaaacca attcgactgt
     1081 tagccaatca ttttcaggtt cagattccta aaatagatgt gtatcactat gatgtggata
     1141 ttaagcctga aaaacggcct cgtagagtca acagggaggt agtagataca atggtgcggc
     1201 acttcaagat gcaaatattt ggtgatcggc agcctgggta tgatggcaaa agaaacatgt
     1261 acacagcaca tccactacca attggacggg atagggttga tatggaggtg actcttccag
     1321 gcgagggtaa agaccaaaca tttaaagtgt ctgttcagtg ggtgtcagtt gtgagccttc
     1381 agttgctttt agaagctttg gctgggcact tgaatgaagt cccagatgac tcagtacaag
     1441 cacttgatgt tatcacaaga caccttccct ccatgaggta caccccagtg ggccgttcct
     1501 ttttctcacc cccggaaggt tactaccacc ctctgggagg gggcagggag gtctggtttg
     1561 gttttcatca gtctgtgaga cctgccatgt ggaatatgat gctcaacatt gatgtatctg
     1621 caactgcttt ctaccgggct cagcctatca ttgagttcat gtgtgaggtt ttagacattc
     1681 agaacatcaa tgaacagacc aaacctctaa cagactccca gcgtgtcaaa tttaccaaag
     1741 aaatcagagg tctcaaagtt gaggtgaccc actgtggaca gatgaaacga aaataccgag
     1801 tttgtaatgt gactagacgg ccagccagtc atcaaacttt tcctttgcag ctagaaaacg
     1861 gtcaagctat ggaatgtaca gtagctcaat attttaagca aaagtatagt ctgcaactga
     1921 aataccccca tcttccctgt ctccaagtgg gacaagaaca aaagcataca tacttgccac
     1981 tcgaggtctg taatatagtg gcaggacagc gatgtatcaa gaagctcaca gacaatcaga
     2041 cttccacaat gatcaaagct acagcaagat ctgctcctga cagacaggaa gagatcagta
     2101 gactggtgaa gagcaacagt atggtgggtg gacctgatcc ataccttaaa gaatttggta
     2161 ttgttgtcca caatgaaatg acagagctca caggcagggt acttccagca ccaatgctgc
     2221 aatatggagg ccggaataaa acagtagcca cacccaacca gggtgtctgg gacatgcgag
     2281 gaaagcagtt ttatgctggc attgaaatta aagtttgggc agttgcttgt tttgcacctc
     2341 agaaacaatg tagggaagat ttactaaaga gtttcactga ccagctgcgt aaaatctcta
     2401 aggatgcagg aatgcccatc cagggtcagc catgtttctg caagtatgca caaggtgcag
     2461 acagtgtgga gcctatgttt aaacatctga aaatgactta tgtgggccta cagctaatag
     2521 tggttatcct gcctggaaag acaccagtat atgcggaggt gaaacgtgtt ggagataccc
     2581 ttctaggtat ggccacacag tgtgtccagg taaaaaatgt agtgaagacc tcacctcaaa
     2641 ccctttccaa tctttgcctg aagataaatg caaaacttgg aggaattaac aatgtgcttg
     2701 tgcctcatca aaggccctcg gtgttccagc agcctgtcat cttcctggga gcggatgtca
     2761 cacacccccc agcaggggat gggaagaaac cttccattgc tgctgtggtt ggcagtatgg
     2821 atggccaccc cagccggtac tgtgccaccg ttcgggtgca gacttcccgg caggagatct
     2881 cccaagagct cctctacagt caagaggtca tccaggacct gactaacatg gttcgagagc
     2941 tgctgattca gttctacaaa tccacacgct tcaaacccac tcggatcatc tattaccgtg
     3001 gaggggtatc tgagggacaa atgaaacagg tagcttggcc agaactaata gcaattcgaa
     3061 aggcatgtat tagcttggaa gaagattacc ggccaggaat aacttatatt gtggtgcaaa
     3121 aaagacatca cacacgactc ttctgtgcag ataaaacaga aagggtaggg aaaagtggca
     3181 atgtaccagc aggcactaca gtggatagta ccatcacaca tccatctgag tttgactttt
     3241 acctctgtag tcatgcagga atacagggaa ccagccgtcc ctcacattac caggtcttgt
     3301 gggatgacaa ctgcttcact gcagatgaac tccagctact gacttaccag ctgtgtcaca
     3361 cctatgtgag gtgcactcgc tcagtctcta ttccagcccc tgcatattat gcccggcttg
     3421 tagcatttag ggcaaggtat catctggtgg ataaagatca tgacagtgcg gaaggcagtc
     3481 atgtgtcagg acagagcaac ggccgagatc ctcaggcctt ggctaaggct gtgcaaatcc
     3541 accatgatac ccagcacacg atgtattttg cctgagaatt cagtggatcc actagtaacg
     3601 gccgccagtg tgctggaatt aattcgctgt ctgcgagggc cagctgttgg ggtgagtact
     3661 ccctctcaaa agcgggcatg acttctgcgc taagattgtc agtttccaaa aacgaggagg
     3721 atttgatatt cacctggccc gcggtgatgc ctttgagggt ggccgcgtcc atctggtcag
     3781 aaaagacaat ctttttgttg tcaagcttga ggtgtggcag gcttgagatc tggccataca
     3841 cttgagtgac aatgacatcc actttgcctt tctctccaca ggtgtccact cccaggtcca
     3901 actgcaggtc gagcatgcat ctagggcggc caattccgcc cctctccctc ccccccccct
     3961 aacgttactg gccgaagccg cttggaataa ggccggtgtg cgtttgtcta tatgtgattt
     4021 tccaccatat tgccgtcttt tggcaatgtg agggcccgga aacctggccc tgtcttcttg
     4081 acgagcattc ctaggggtct ttcccctctc gccaaaggaa tgcaaggtct gttgaatgtc
     4141 gtgaaggaag cagttcctct ggaagcttct tgaagacaaa caacgtctgt agcgaccctt
     4201 tgcaggcagc ggaacccccc acctggcgac aggtgcctct gcggccaaaa gccacgtgta
     4261 taagatacac ctgcaaaggc ggcacaaccc cagtgccacg ttgtgagttg gatagttgtg
     4321 gaaagagtca aatggctctc ctcaagcgta ttcaacaagg ggctgaagga tgcccagaag
     4381 gtaccccatt gtatgggatc tgatctgggg cctcggtgca catgctttac atgtgtttag
     4441 tcgaggttaa aaaaacgtct aggccccccg aaccacgggg acgtggtttt cctttgaaaa
     4501 acacgatgat aagcttgcca caacccggga taattcctgc agccaatatg ggatcggcca
     4561 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     4621 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     4681 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg
     4741 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     4801 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     4861 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     4921 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     4981 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     5041 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg
     5101 atgatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     5161 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     5221 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     5281 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     5341 agttcttctg aggggatcaa ttctctagag ctcgctgatc agcctcgact gtgccttcta
     5401 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     5461 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     5521 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     5581 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg
     5641 gctcgagtgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     5701 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
     5761 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     5821 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     5881 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     5941 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     6001 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     6061 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     6121 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     6181 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     6241 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     6301 gcctttctcc cttcgggaag cgtggcgctt tctcaatgct cacgctgtag gtatctcagt
     6361 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     6421 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     6481 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     6541 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     6601 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     6661 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     6721 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     6781 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     6841 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     6901 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     6961 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     7021 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     7081 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     7141 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     7201 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     7261 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     7321 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     7381 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     7441 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     7501 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     7561 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     7621 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     7681 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     7741 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     7801 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     7861 ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
