LOCUS       sequence-228916-        8266 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 8266)
  AUTHORS   Lian SL, Li S, Abadal GX, Pauley BA, Fritzler MJ, Chan EK
  TITLE     The C-terminal half of human Ago2 binds to multiple GW-rich regions 
            of GW182 and requires GW182 to mediate silencing.
  JOURNAL   RNA. 2009 May . 15(5):804-13.
  PUBMED    19324964
REFERENCE   2  (bases 1 to 8266)
  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: "RNA. 2009 
            May . 15(5):804-13."
COMMENT     Sequence Label: GFP-Ago4
FEATURES             Location/Qualifiers
     source          1..8266
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     953..972
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             join(999..1001,1002..1004,1005..1715)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKAEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1044..1065)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1305..1324)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1652..1673
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    1731..1755
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      1917..1926
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(5261..5285)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    5322..5456
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5346..5365)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5400..5419
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5572..5588)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5572..5588)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5585..5607)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5596..5612
                     /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(5620..5626,5627..5644,5645..5650))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5665..5686
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5803..5820)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5974..6562)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6054..6073)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6733..7524,7525..7593))
                     /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     7356..7375
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7594..7698)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7766..7784
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7822..7844)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7944..7963
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8157..8179
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8171..8188
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     8172..8188
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatctgaat tcctgttggg ctcgcggttg aggacaaact cttcgcggtc tttccagtac
      661 tcttggatcg gaaacccgtc ggcctccgaa cggtactccg ccaccgaggg acctgagcga
      721 gtccgcatcg accggatcgg aaaacctctc gagaaaggcg tctaaccagt cacagtcgca
      781 aggtaggctg agcaccgtgg cgggcggcag cgggtggcgg tcggggttgt ttctggcgga
      841 ggtgctgctg atgatgtaat taaagtaggc ggtcttgaga cggcggatgg tcgaggtgag
      901 gtgtggcagg cttgagatct ggccatacac ttgagtgaca atgacatcca ctttgccttt
      961 ctctccacag gtgtccactc ccaggtccaa ccggatctat ggtgagcaag gccgaggagc
     1021 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     1081 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
     1141 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
     1201 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     1261 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
     1321 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     1381 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca
     1441 gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga
     1501 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     1561 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc
     1621 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     1681 ccgggatcac tctcggcatg gacgagctgt acaagtctag aggatcatca acaagtttgt
     1741 acaaaaaagc aggctttggg ccgggcgccg ccgccgcccc ctgcccagcg cccgcgtctc
     1801 cgcggcgcca ccccagcgcc aatattccgg agatcaagcg ttacgcggcg gcggcggcgg
     1861 cggcggcggg gcccggagcg ggaggcgccg gggaccgggg cgaggcggcc cccgccgccg
     1921 ccatggaggc gctgggaccc ggacctccgg ctagcctgtt tcagccacct cgtcgtcctg
     1981 gccttggaac tgttggaaaa ccaattcgac tgttagccaa tcattttcag gttcagattc
     2041 ctaaaataga tgtgtatcac tatgatgtgg atattaagcc tgaaaaacgg cctcgtagag
     2101 tcaacaggga ggtagtagat acaatggtgc ggcacttcaa gatgcaaata tttggtgatc
     2161 ggcagcctgg gtatgatggc aaaagaaaca tgtacacagc acatccacta ccaattggac
     2221 gggatagggt tgatatggag gtgactcttc caggcgaggg taaagaccaa acatttaaag
     2281 tgtctgttca gtgggtgtca gttgtgagcc ttcagttgct tttagaagct ttggctgggc
     2341 acttgaatga agtcccagat gactcagtac aagcacttga tgttatcaca agacaccttc
     2401 cctccatgag gtacacccca gtgggccgtt cctttttctc acccccggaa ggttactacc
     2461 accctctggg agggggcagg gaggtctggt ttggttttca tcagtctgtg agacctgcca
     2521 tgtggaatat gatgctcaac attgatgtat ctgcaactgc tttctaccgg gctcagccta
     2581 tcattgagtt catgtgtgag gttttagaca ttcagaacat caatgaacag accaaacctc
     2641 taacagactc ccagcgtgtc aaatttacca aagaaatcag aggtctcaaa gttgaggtga
     2701 cccactgtgg acagatgaaa cgaaaatacc gagtttgtaa tgtgactaga cggccagcca
     2761 gtcatcaaac ttttcctttg cagctagaaa acggtcaagc tatggaatgt acagtagctc
     2821 aatattttaa gcaaaagtat agtctgcaac tgaaataccc ccatcttccc tgtctccaag
     2881 tgggacaaga acaaaagcat acatacttgc cactcgaggt ctgtaatata gtggcaggac
     2941 agcgatgtat caagaagctc acagacaatc agacttccac aatgatcaaa gctacagcaa
     3001 gatctgctcc tgacagacag gaagagatca gtagactggt gaagagcaac agtatggtgg
     3061 gtggacctga tccatacctt aaagaatttg gtattgttgt ccacaatgaa atgacagagc
     3121 tcacaggcag ggtacttcca gcaccaatgc tgcaatatgg aggccggaat aaaacagtag
     3181 ccacacccaa ccagggtgtc tgggacatgc gaggaaagca gttttatgct ggcattgaaa
     3241 ttaaagtttg ggcagttgct tgttttgcac ctcagaaaca atgtagggaa gatttactaa
     3301 agagtttcac tgaccagctg cgtaaaatct ctaaggatgc aggaatgccc atccagggtc
     3361 agccatgttt ctgcaagtat gcacaaggtg cagacagtgt ggagcctatg tttaaacatc
     3421 tgaaaatgac ttatgtgggc ctacagctaa tagtggttat cctgcctgga aagacaccag
     3481 tatatgcgga ggtgaaacgt gttggagata cccttctagg tatggccaca cagtgtgtcc
     3541 aggtaaaaaa tgtagtgaag acctcacctc aaaccctttc caatctttgc ctgaagataa
     3601 atgcaaaact tggaggaatt aacaatgtgc ttgtgcctca tcaaaggccc tcggtgttcc
     3661 agcagcctgt catcttcctg ggagcggatg tcacacaccc cccagcaggg gatgggaaga
     3721 aaccttccat tgctgctgtg gttggcagta tggatggcca ccccagccgg tactgtgcca
     3781 ccgttcgggt gcagacttcc cggcaggaga tctcccaaga gctcctctac agtcaagagg
     3841 tcatccagga cctgactaac atggttcgag agctgctgat tcagttctac aaatccacac
     3901 gcttcaaacc cactcggatc atctattacc gtggaggggt atctgaggga caaatgaaac
     3961 aggtagcttg gccagaacta atagcaattc gaaaggcatg tattagcttg gaagaagatt
     4021 accggccagg aataacttat attgtggtgc aaaaaagaca tcacacacga ctcttctgtg
     4081 cagataaaac agaaagggta gggaaaagtg gcaatgtacc agcaggcact acagtggata
     4141 gtaccatcac acatccatct gagtttgact tttacctctg tagtcatgca ggaattcagg
     4201 gaaccagccg tccctcacat taccaggtct tgtgggatga caactgcttc actgcagatg
     4261 aactccagct actgacttac cagctgtgtc acacctatgt gaggtgcact cgctcagtct
     4321 ctattccagc ccctgcatat tatgcccggc ttgtagcatt tagggcaagg tatcatctgg
     4381 tggataaaga tcatgacagt gcggaaggca gtcatgtgtc aggacagagc aacggccggg
     4441 atcctcaggc cttggctaag gctgtgcaaa tccaccatga tacccagcac acgatgtatt
     4501 ttgcctgaga gtctcagaaa aagaactcaa ccaatttggc accccatgca gcctcaaaat
     4561 gtttcaaatg cctaccgcct ctagatcgag ccacgttgac ttcaggtggt cttctaccag
     4621 cagctcggaa tagttgcact gaatctatac tttgcagcac tgtctgatgc gtcaacaaaa
     4681 ttgagccatt tttttaaagt aatagatact aatagattat cttttctgat gcactggact
     4741 gaatttttac ctcaatgtgc aaaggctgat cagcctgaac tttctaaagg actttacaag
     4801 aagggtttct atggaatatt ttgctagctg tggtgttcac cgcatccctc tagtcttaga
     4861 agagaagatt attccttttt tctgtagtca ttgagtgggg tatatgcata agtgggagag
     4921 aaaaaccaaa caatctactt cagttcagtg taactattta attgtgtggg tccatagact
     4981 ttttaatgaa gatttctgac tttgccactg taaatgcctt taatgagcat taatttttga
     5041 aagttttaca gagttttatt agttttacta ctttatctta ttgatctctg cagacttgtg
     5101 ctggtgcaag tacatgctga caggcaattt gcactatatt tggctgcaga ttcaaatagg
     5161 cagttctctt atttggtcga cttttgtgaa tgtgtgacat aaacagatgt aatgcatgtc
     5221 acagtgacgg agataacact gccatgataa aagtatctag acccagcttt cttgtacaaa
     5281 gtggttgatt cgacctcgag gccttaagtg actaggtacc cgatccagac atgataagat
     5341 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
     5401 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     5461 acaacaattg cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaaa
     5521 gcaagtaaaa cccaagctgg gaagctcaga tctgagcttg gcgtaatcat ggtcatagct
     5581 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     5641 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     5701 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     5761 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     5821 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     5881 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5941 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     6001 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     6061 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     6121 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     6181 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     6241 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     6301 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     6361 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     6421 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     6481 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6541 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6601 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     6661 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6721 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6781 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6841 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6901 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6961 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     7021 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     7081 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     7141 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     7201 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     7261 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     7321 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     7381 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     7441 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     7501 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     7561 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     7621 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     7681 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     7741 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg
     7801 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     7861 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     7921 gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat
     7981 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc
     8041 attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca
     8101 gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca
     8161 gtcacgacgt tgtaaaacga cggccagtga attccattgc atacgttgta tctatatcat
     8221 aatatgtaca tttatattgg ctcatgtcca atatgaccgc catgtt
//
