LOCUS       sequence-234052-        6907 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  dsRNA expression .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6907)
  TITLE     Petr Svoboda lab unpublished
REFERENCE   2  (bases 1 to 6907)
  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"
COMMENT     Sequence Label: CAG-EGFP-Lin28IR
FEATURES             Location/Qualifiers
     source          1..6907
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..662
                     /label=chicken beta-actin promoter
     intron          663..1679
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1603..1625
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1687..1706
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1734..1743
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1740..1742,1743..1745,1746..2459)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1740..1742,1743..1745,1746..2456)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1785..1806)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2046..2065)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2393..2414
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3929..3948)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3994..4049
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4048..4067)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4410..4426)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4410..4426)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4423..4445)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4434..4450
                     /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(4458..4464,4465..4482,4483..4488))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4503..4524
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4583..4778
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4629..4764
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4784..4918
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4821..4840)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4875..4894
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4986..5003)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5157..5745)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5237..5256)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5916..6707,6708..6776))
                     /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     6539..6558
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6777..6881)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggnsgcg cgccrggsgg ggsggggsgg ggsgrggggs
      541 ggggsggggc gaggcggaga ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt
      601 ttccttttat ggcgaggcgg cggcggcggc ggccctataa aaagcgaagc gcgcggcggg
      661 cgggagtcgc tgcgcgctgc cttcgccccg tgccccgctc cgccgccgcc tcgcgccgcc
      721 cgccccggct ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc
      781 ctccgggctg taattagcgc ttggtttaat gacggcttgt ttcttttctg tggctgcgtg
      841 aaagccttga ggggctccgg gagggccctt tgtgcggggg gagcggctcg gggggtgcgt
      901 gcgtgtgtgt gtgcgtgggg agcgccgcgt gcggctccgc gctgcccggc ggctgtgagc
      961 gctgcgggcg cggcgcgggg ctttgtgcgc tccgcagtgt gcgcgagggg agcgcggccg
     1021 ggggcggtgc cccgcggtgc ggggggggct gcgaggggaa caaaggctgc gtgcggggtg
     1081 tgtgcgtggg ggggtgagca gggggtgtgg gcgcgtcggt cgggctgcaa ccccccctgc
     1141 acccccctcc ccgagttgct gagcacggcc cggcttcggg tgcggggctc cgtacggggc
     1201 gtggcgcggg gctcgccgtg ccgggcgggg ggtggcggca ggtgggggtg ccgggcgggg
     1261 cggggccgcc tcgggccggg gagggctcgg gggaggggcg cggcggcccc cggagcgccg
     1321 gcggctgtcg aggcgcggcg agccgcagcc attgcctttt atggtaatcg tgcgagaggg
     1381 cgcagggact tcctttgtcc caaatctgtg cggagccgaa atctgggagg cgccgccgca
     1441 ccccctctag cgggcgcggg gcgaagcggt gcggcgccgg caggaaggaa atgggcgggg
     1501 agggccttcg tgcgtcgccg cgccgccgtc cccttctccc tctccagcct cggggctgtc
     1561 cgcgggggga cggctgcctt cgggggggac ggggcagggc ggggttcggc ttctggcgtg
     1621 tgaccggcgg ctctagagcc tctgctaacc atgttcatgc cttcttcttt ttcctacagc
     1681 tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt tggcaaagaa ttcgccacca
     1741 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     1801 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
     1861 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     1921 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
     1981 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     2041 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     2101 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     2161 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     2221 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     2281 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     2341 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     2401 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtaag
     2461 aattcactcc tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg gccaatgccc
     2521 tggctcgaga tagctagctt cgcggccgcg actctagagt gctgtactct tatgtgaaat
     2581 cctcacacat acatacagat tcaagtctcc cctctcactt atgcacatac tatatgtagt
     2641 atctcaattc tataaaacat gacaccctgc ttaaatgtag tttgcttaag agtgatttgt
     2701 ctaattaaga acttcccagt tcctcagcag tagttaacat cccggccggc tgctctctgt
     2761 ggaccactat tcccctgtgc ttgcatgagg tagacttccc gcccgggtga agggaacact
     2821 gacaagtact aagtcttttt ccgtttctga atcaaaggaa gcttggtttg acacttgttt
     2881 cgcttcccgt ctggcaaggg aaatatcaca cagccttcta caaaagaact tccctagccc
     2941 agccacttgg caaagcctcg ggttctaggg tggttcctcc ccacaaaagc cttcacttgc
     3001 tttcagatag agtgtggtag gagttccttt gtctcatcct ttccttccct ccaaggaaag
     3061 acagggtgac actgggaacc ccggcctgag acacggcagt gcaagctacc cacgcaaccc
     3121 acccttaccc ccactttctc cactctgcag attgatgcct ctttccttaa actccccatt
     3181 agccaaagaa taaccctgac tcctgggcct caattctggg cttctgggag cagggcaggg
     3241 ctgtggatct cttcctcttc ctccctggat ccaggagttt aaggaaagag gcatcaatct
     3301 gcagagtgga gaaagtgggg gtaagggtgg gttgcgtggg tagcttgcac tgccgtgtct
     3361 caggccgggg ttcccagtgt caccctgtct ttccttggag ggaaggaaag gatgagacaa
     3421 aggaactcct accacactct atctgaaagc aagtgaaggc ttttgtgggg aggaaccacc
     3481 ctagaacccg aggctttgcc aagtggctgg gctagggaag ttcttttgta gaaggctgtg
     3541 tgatatttcc cttgccagac gggaagcgaa acaagtgtca aaccaagctt cctttgattc
     3601 agaaacggaa aaagacttag tacttgtcag tgttcccttc acccgggcgg gaagtctacc
     3661 tcatgcaagc acaggggaat agtggtccac agagagcagc cggccgggat gttaactact
     3721 gctgaggaac tgggaagttc ttaattagac aaatcactct taagcaaact acatttaagc
     3781 agggtgtcat gttttataga attgagatac tacatatagt atgtgcataa gtgagagggg
     3841 agacttgaat ctgtatgtat gtgtgaggat ttcacataag agtacaggca ctctagagcg
     3901 gccgcgttgg taccaaccat ggtgtagatc tttttccctc tgccaaaaat tatggggaca
     3961 tcatgaagcc ccttgagcat ctgacttctg gctaataaag gaaatttatt ttcattgcaa
     4021 tagtgtgttg gaattttttg tgtctctcac tcggaaggac atatgggagg gcaaatcatt
     4081 taaaacatca gaatgagtat ttggtttaga gtttggcaac atatgcccat atgctggctg
     4141 ccatgaacaa aggttggcta taaagaggtc atcagtatat gaaacagccc cctgctgtcc
     4201 attccttatt ccatagaaaa gccttgactt gaggttagat tttttttata ttttgttttg
     4261 tgttattttt ttctttaaca tccctaaaat tttccttaca tgttttacta gccagatttt
     4321 tcctcctctc ctgactactc ccagtcatag ctgtccctct tctcttatgg agatccctcg
     4381 acctgcagcc caagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     4441 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     4501 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4561 aacctgtcgt gccagcggat ccgcatctca attagtcagc aaccatagtc ccgcccctaa
     4621 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     4681 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     4741 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctaacttgt ttattgcagc
     4801 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     4861 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctggatccg
     4921 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     4981 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     5041 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     5101 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     5161 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     5221 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     5281 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     5341 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     5401 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     5461 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     5521 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     5581 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     5641 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     5701 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     5761 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     5821 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     5881 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     5941 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     6001 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     6061 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     6121 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     6181 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     6241 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     6301 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     6361 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     6421 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     6481 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     6541 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     6601 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     6661 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     6721 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     6781 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     6841 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     6901 ccacctg
//
