LOCUS       sequence-379201-        5291 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For mammalian expression of T1 and AP3, the RNA aptamers of ncOGT 
            and GFP, respectively. Expression of each aptamer is driven by a U6 
            promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5291)
  AUTHORS   Zhu Y, Hart GW
  TITLE     Dual-specificity RNA aptamers enable manipulation of target-specific
            O-GlcNAcylation and unveil functions of O-GlcNAc on beta-catenin.
  JOURNAL   Cell. 2022 Dec 30:S0092-8674(22)01529-X. doi: 
            10.1016/j.cell.2022.12.016.
  PUBMED    36626902
REFERENCE   2  (bases 1 to 5291)
  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: "Cell. 2022 
            Dec 30:S0092-8674(22)01529-X. doi: 10.1016/j.cell.2022.12.016."
COMMENT     Sequence Label: pSilencer2.1-U6-T1-U6-AP3
FEATURES             Location/Qualifiers
     source          1..5291
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        262..472
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        284..480
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      331..466
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     393..412
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             529..1554
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             529..1554
                     /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    1684..1805
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1721..1740)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1775..1794
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1854..1870)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1878..1894
                     /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(1956..2024,2025..2816)
                     /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(2174..2193)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2987..3575
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3476..3495
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3729..3746
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3863..3884
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3899..3904,3905..3922,3923..3929)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3937..3953
                     /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     3961..3977
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3997..4015
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3997..4014
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     4027..4046
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4027..4045
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        4176..4424
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(4542..4559)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4542..4558)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4551..4573)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        4640..4888
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(5122..5141)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5241..5263
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattccag caggcagaag tatgcaaagc atgcatctca attagttagc aaccaggtgt
      241 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
      301 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
      361 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct
      421 gcctctgagc tattccagaa gtagtgggga ggcttttttg gaggcctagg cttttgcaaa
      481 aagctcccgg gagcttgtat atccattttc ggatctgatc agcacgtgat gaaaaagcct
      541 gaactcaccg cgacgtctgt cgagaagttt ctgatcgaaa agttcgacag cgtctccgac
      601 ctgatgcagc tctcggaggg cgaagaatct cgtgctttca gcttcgatgt aggagggcgt
      661 ggatatgtcc tgcgggtaaa tagctgcgcc gatggtttct acaaagatcg ttatgtttat
      721 cggcactttg catcggccgc gctcccgatt ccggaagtgc ttgacattgg ggaattcagc
      781 gagagcctga cctattgcat ctcccgccgt gcacagggtg tcacgttgca agacctgcct
      841 gaaaccgaac tgcccgctgt tctgcagccg gtcgcggagg ccatggatgc gatcgctgcg
      901 gccgatctta gccagacgag cgggttcggc ccattcggac cgcaaggaat cggtcaatac
      961 actacatggc gtgatttcat atgcgcgatt gctgatcccc atgtgtatca ctggcaaact
     1021 gtgatggacg acaccgtcag tgcgtccgtc gcgcaggctc tcgatgagct gatgctttgg
     1081 gccgaggact gccccgaagt ccggcacctc gtgcacgcgg atttcggctc caacaatgtc
     1141 ctgacggaca atggccgcat aacagcggtc attgactgga gcgaggcgat gttcggggat
     1201 tcccaatacg aggtcgccaa catcttcttc tggaggccgt ggttggcttg tatggagcag
     1261 cagacgcgct acttcgagcg gaggcatccg gagcttgcag gatcgccgcg gctccgggcg
     1321 tatatgctcc gcattggtct tgaccaactc tatcagagct tggttgacgg caatttcgat
     1381 gatgcagctt gggcgcaggg tcgatgcgac gcaatcgtcc gatccggagc cgggactgtc
     1441 gggcgtacac aaatcgcccg cagaagcgcg gccgtctgga ccgatggctg tgtagaagta
     1501 ctcgccgata gtggaaaccg acgccccagc actcgtccga gggcaaagga atagcacgtg
     1561 ctacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     1621 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     1681 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     1741 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     1801 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag
     1861 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     1921 ataaaggtaa gcctgaatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     1981 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     2041 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     2101 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     2161 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     2221 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     2281 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     2341 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     2401 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     2461 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     2521 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     2581 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     2641 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     2701 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     2761 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     2821 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     2881 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     2941 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     3001 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttttt
     3061 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     3121 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     3181 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     3241 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     3301 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     3361 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     3421 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     3481 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     3541 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     3601 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     3661 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     3721 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc
     3781 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
     3841 cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt
     3901 tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca
     3961 caggaaacag ctatgacatg attacgaatt gcaacgattt aggtgacact atagaagaga
     4021 aggaattaat acgactcact atagggagag agagagaatt accctcacta aagggaggag
     4081 aagcatgaat tccccagtgg aaagacgcgc aggcaaaacg caccacgtga cggagcgtga
     4141 ccgcgcgccg agcgcgcgcc aaggtcgggc aggaagaggg cctatttccc atgattcctt
     4201 catatttgca tatacgatac aaggctgtta gagagataat tagaattaat ttgactgtaa
     4261 acacaaagat attagtacaa aatacgtgac gtagaaagta ataatttctt gggtagtttg
     4321 cagttttaaa attatgtttt aaaatggact atcatatgct taccgtaact tgaaagtatt
     4381 tcgatttctt gggtttatat atcttgtgga aaggacgaaa caccgggaga atcacttacg
     4441 gatcctcgtt aagtggctct cagtcgcccg taactccgat gccgatgcag cggcgacaac
     4501 tgcacccaag gctcggaagc ttttttttgg aaaagcttgg cactggccgt cgttttacaa
     4561 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     4621 ttccaaggtc gggcaggaag agggcctatt tcccatgatt ccttcatatt tgcatatacg
     4681 atacaaggct gttagagaga taattagaat taatttgact gtaaacacaa agatattagt
     4741 acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt taaaattatg
     4801 ttttaaaatg gactatcata tgcttaccgt aacttgaaag tatttcgatt tcttgggttt
     4861 atatatcttg tggaaaggac gcgggatcag cttctggact gcgatgggag cacgaaacgt
     4921 cgtggcgcaa ttgggtgggg aaagtcctta aaagagggcc accacagaag ctttttttgc
     4981 cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct
     5041 gaatggcgaa tggcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca
     5101 ccgcatatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     5161 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     5221 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     5281 gaaacgcgcg a
//
