LOCUS       sequence-238931-        6643 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  siRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6643)
  TITLE     Oka Lab plasmids
REFERENCE   2  (bases 1 to 6643)
  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: AV5-siRNA-GFP
FEATURES             Location/Qualifiers
     source          1..6643
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   69..198
                     /label=AAV2 ITR
     promoter        426..739
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     658..678
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     primer_bind     755..771
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     756..772
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(814..847)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     enhancer        963..1266
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1267..1470
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1421..1441
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1509..1518
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1515..1517,1518..1520,1521..2234)
                     /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(1515..1517,1518..1520,1521..2231)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1560..1581)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1821..1840)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2168..2189
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2248..2836
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2301..2321)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2719..2730)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2844..3466
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3043..3062)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     promoter        complement(3768..3786)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3769..3786)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3804..3820)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3804..3820)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3817..3839)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3828..3844
                     /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(3852..3858,3859..3876,3877..3882))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3897..3918
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4206..4794)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4286..4305)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4965..5756,5757..5825))
                     /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     5588..5607
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5826..5930)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6008..6463)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6145..6166)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6357..6376
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6589..6611
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6603..6620
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6604..6620
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     join(6627..6643,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(6627..6643,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gggcgaattg ggcccggcgc gccgacgtcg catgctcctc tagactcgat cgaggaattc
       61 ggtaccccct gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg
      121 cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact
      181 ccatcactag gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc
      241 catgctctag agtggagtcg tgacgtgaat tacgtcatag ggttagggag gtcctgtatt
      301 agaggtcacg tgagtgtttt gcgacatttt gcgacaccat gtggtcacgc tgggtattta
      361 agcccgagtg agcacgcagg gtctccatac gcgccgtcga cgagctcaga tctccccgct
      421 ctagagatcc gacgccgcca tctctaggcc cgcgccggcc ccctcgcaca gacttgtggg
      481 agaagctcgg ctactcccct gccccggtta atttgcatat aatatttcct agtaactata
      541 gaggcttaat gtgcgataaa agacagataa tctgttcttt ttaatactag ctacatttta
      601 catgataggc ttggatttct ataagagata caaatactaa attattattt taaaaaacag
      661 cacaaaagga aactcaccct aactgtaaag taattgtgtg ttttgagact ataaatatcc
      721 cttggagaaa agccttgtta acgcgcggtg accctcgagg tcgacggtat cgataagctc
      781 gcttcacgag attccagcag gtcgagggac ctaataactt cgtatagcat acattatacg
      841 aagttatatt aagggttcca agcttaagcg gccgcgtgga taaccgtatt accgccatgc
      901 attagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
      961 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
     1021 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
     1081 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
     1141 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
     1201 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
     1261 accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
     1321 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
     1381 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
     1441 gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat ccgctagcgc
     1501 taccggtcgc caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc
     1561 tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg
     1621 gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg
     1681 tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc
     1741 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
     1801 agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg
     1861 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
     1921 acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg
     1981 acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca
     2041 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     2101 tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc
     2161 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     2221 agctgtacaa gtaggaattc tatcgataat caacctctgg attacaaaat ttgtgaaaga
     2281 ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg
     2341 cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc
     2401 tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc
     2461 actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt
     2521 tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt
     2581 gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg
     2641 aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg
     2701 tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg
     2761 ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt
     2821 tgggccgcct ccccgcatcg atagggtggc atccctgtga cccctcccca gtgcctctcc
     2881 tggccctgga agttgccact ccagtgccca ccagccttgt cctaataaaa ttaagttgca
     2941 tcattttgtc tgactaggtg tccttctata atattatggg gtggaggggg gtggtatgga
     3001 gcaaggggca agttgggaag acaacctgta gggcctgcgg ggtctattgg gaaccaagct
     3061 ggagtgcagt ggcacaatct tggctcactg caatctccgc ctcctgggtt caagcgattc
     3121 tcctgcctca gcctcccgag ttgttgggat tccaggcatg catgaccagg ctcagctaat
     3181 ttttgttttt ttggtagaga cggggtttca ccatattggc caggctggtc tccaactcct
     3241 aatctcaggt gatctaccca ccttggcctc ccaaattgct gggattacag gcgtgaacca
     3301 ctgctccctt ccctgtcctt ctgattttaa aataactata ccagcaggag gacgtccaga
     3361 cacagcatag gctacctggc catgcccaac cggtgggaca tttgagttgc ttgcttggca
     3421 ctgtcctctc atgcgttggg tccactcagt agatgcctgt tgaattgggc tgcagacgcg
     3481 tgtctagagg agcatgcgcg tcggcgcgtc gctctagagc atggctacgt agataagtag
     3541 catggcgggt taatcattaa ctacaaggaa cccctagtga tggagttggc cactccctct
     3601 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt
     3661 gcccgggcgg cctcagtgag cgagcgagcg cgccaggggt tcgaaatcga taagcttgga
     3721 tcgatccgga gagctcgtcg acggcgcgcc atgcatagct tgagtattct atagtgtcac
     3781 ctaaatagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     3841 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     3901 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     3961 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     4021 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     4081 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     4141 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     4201 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     4261 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     4321 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     4381 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     4441 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     4501 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     4561 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     4621 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     4681 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     4741 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     4801 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     4861 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     4921 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     4981 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     5041 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     5101 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     5161 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     5221 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     5281 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     5341 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     5401 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     5461 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     5521 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     5581 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     5641 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     5701 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     5761 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     5821 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     5881 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     5941 cgaaaagtgc cacctgatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
     6001 catcaggaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     6061 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     6121 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     6181 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     6241 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     6301 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     6361 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     6421 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtccattcg ccattcaggc
     6481 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
     6541 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
     6601 gttgtaaaac gacggccagt gaattgtaat acgactcact ata
//
