LOCUS       sequence-379580-        4887 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of a siRNA that targets Opa1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4887)
  AUTHORS   Scorrano L
  TITLE     Luca Scorrano plasmids
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 4887)
  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: 
            "unpublished"
COMMENT     Sequence Label: pSilencer-Opa1 KO
FEATURES             Location/Qualifiers
     source          1..4887
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      171..759
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     660..679
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     913..930
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1047..1068
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1083..1088,1089..1106,1107..1113)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1121..1137
                     /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)."
     primer_bind     1145..1161
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1181..1199
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1181..1198
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     1211..1230
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1211..1229
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        1360..1608
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1360..1380
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1531..1550
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(1677..1694)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1677..1693)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1686..1708)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1902..1921)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2021..2043
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2081..2099)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2333..2543
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        2355..2551
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2402..2537
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2464..2483
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2600..3625
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             2600..3625
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    3755..3876
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3792..3811)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3846..3865
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3925..3941)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3949..3965
                     /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)."
     CDS             join(4027..4095,4096..4887)
                     /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     complement(4245..4264)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
       61 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
      121 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
      181 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
      241 tttttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
      301 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
      361 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
      421 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
      481 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
      541 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
      601 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
      661 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
      721 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
      781 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
      841 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
      901 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg
      961 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
     1021 aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag
     1081 gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt
     1141 cacacaggaa acagctatga catgattacg aattgcaacg atttaggtga cactatagaa
     1201 gagaaggaat taatacgact cactataggg agagagagag aattaccctc actaaaggga
     1261 ggagaagcat gaattcccca gtggaaagac gcgcaggcaa aacgcaccac gtgacggagc
     1321 gtgaccgcgc gccgagcgcg cgccaaggtc gggcaggaag agggcctatt tcccatgatt
     1381 ccttcatatt tgcatatacg atacaaggct gttagagaga taattagaat taatttgact
     1441 gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag
     1501 tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag
     1561 tatttcgatt tcttgggttt atatatcttg tggaaaggac gcgggatccc gtcatcagtc
     1621 tgagccaggt tcaagagacc tggctcagac tgatgacttt tttggaaaag cttggcactg
     1681 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     1741 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     1801 tcccaacagt tgcgcagcct gaatggcgaa tggcgcctga tgcggtattt tctccttacg
     1861 catctgtgcg gtatttcaca ccgcatatgg tgcactctca gtacaatctg ctctgatgcc
     1921 gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     1981 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     2041 aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt
     2101 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga
     2161 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     2221 atgagacaat aaccctgata aatgcttcaa taatattcca gcaggcagaa gtatgcaaag
     2281 catgcatctc aattagttag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     2341 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     2401 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     2461 ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgggg
     2521 aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt
     2581 cggatctgat cagcacgtga tgaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt
     2641 tctgatcgaa aagttcgaca gcgtctccga cctgatgcag ctctcggagg gcgaagaatc
     2701 tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa atagctgcgc
     2761 cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatcggccg cgctcccgat
     2821 tccggaagtg cttgacattg gggaattcag cgagagcctg acctattgca tctcccgccg
     2881 tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc
     2941 ggtcgcggag gccatggatg cgatcgctgc ggccgatctt agccagacga gcgggttcgg
     3001 cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca tatgcgcgat
     3061 tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca gtgcgtccgt
     3121 cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag tccggcacct
     3181 cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca taacagcggt
     3241 cattgactgg agcgaggcga tgttcgggga ttcccaatac gaggtcgcca acatcttctt
     3301 ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc ggaggcatcc
     3361 ggagcttgca ggatcgccgc ggctccgggc gtatatgctc cgcattggtc ttgaccaact
     3421 ctatcagagc ttggttgacg gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga
     3481 cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc
     3541 ggccgtctgg accgatggct gtgtagaagt actcgccgat agtggaaacc gacgccccag
     3601 cactcgtccg agggcaaagg aatagcacgt gctacgagat ttcgattcca ccgccgcctt
     3661 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     3721 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
     3781 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     3841 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc
     3901 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     3961 cacaattcca cacaacatac gagccggaag cataaaggta agcctgaata ttgaaaaagg
     4021 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     4081 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     4141 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     4201 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     4261 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     4321 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     4381 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     4441 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     4501 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     4561 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     4621 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     4681 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     4741 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     4801 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     4861 ggtgcctcac tgattaagca ttggtaa
//
