LOCUS       sequence-308573-        7086 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tet-repressor with neomycin selection marker, to be used together 
            with pT2-CMV/TetO2-EYFP-GW .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7086)
  AUTHORS   Laidou S, Alanis-Lobato G, Pribyl J, Rasko T, Tichy B, Mikulasek K, 
            Tsagiopoulou M, Oppelt J, Kastrinaki G, Lefaki M, Singh M, Zink A, 
            Chondrogianni N, Psomopoulos F, Prigione A, Ivics Z, Pospisilova S, 
            Skladal P, Izsvak Z, Andrade-Navarro MA, Petrakis S
  TITLE     Nuclear inclusions of pathogenic ataxin-1 induce oxidative stress 
            and perturb the protein synthesis machinery.
  JOURNAL   Redox Biol. 2020 Feb 11;32:101458. doi: 10.1016/j.redox.2020.101458.
  PUBMED    32145456
REFERENCE   2  (bases 1 to 7086)
  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: "Redox 
            Biol."; date: "2020-02-11"; volume: "32:101458. doi"; pages: " 
            10.1016/j.redox.2020.101458"
COMMENT     Sequence Label: pT2-TetR-neoR
FEATURES             Location/Qualifiers
     source          1..7086
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     34..56
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     48..65
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     49..65
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     75..94
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        75..93
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    471..605
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(495..514)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     549..568
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(779..1573)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(890..909)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1500..1519
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1640..1969)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1654..1789
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2082..2306
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2295..2312
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(2332..2955)
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     2863..2882
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(2985..3004)
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     intron          3013..3585
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     promoter        complement(3614..3943)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3628..3763
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4347..4367)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4347..4365)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4386..4402)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4386..4402)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4399..4421)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4410..4426
                     /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(4434..4440,4441..4458,4459..4464))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4479..4500
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4617..4634)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4788..5376)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4868..4887)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5547..6338,6339..6407))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAAKLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6170..6189
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6408..6512)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6539..6994
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6626..6645)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6836..6857
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg
       61 ccagtgagcg cgcgtaatac gactcactat agggcgaatt ggagctcgga tccctataca
      121 gttgaagtcg gaagtttaca tacacttaag ttggagtcat taaaactcgt ttttcaacta
      181 ctccacaaat ttcttgttaa caaacaatag ttttggcaag tcagttagga catctacttt
      241 gtgcatgaca caagtcattt ttccaacaat tgtttacaga cagattattt cacttataat
      301 tcactgtatc acaattccag tgggtcagaa gtttacatac actaagttga ctgtgccttt
      361 aaacagcttg gaaaattcca gaaaatgatg tcatggcttt agaagcttct gatagactaa
      421 ttgacatcat ttgagtcaat tggaggtgta cctgtggatg tatttcaagg gatccagaca
      481 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
      541 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
      601 aagttggggt gggcgaagaa ctccagcatg agatccccgc gctggaggat catccagccg
      661 gcgtcccgga aaacgattcc gaagcccaac ctttcataga aggcggcggt ggaatcgaaa
      721 tctcgtgatg gcaggttggg cgtcgcttgg tcggtcattt cgaaccccag agtcccgctc
      781 agaagaactc gtcaagaagg cgatagaagg cgatgcgctg cgaatcggga gcggcgatac
      841 cgtaaagcac gaggaagcgg tcagcccatt cgccgccaag ctcttcagca atatcacggg
      901 tagccaacgc tatgtcctga tagcggtccg ccacacccag ccggccacag tcgatgaatc
      961 cagaaaagcg gccattttcc accatgatat tcggcaagca ggcatcgcca tgggtcacga
     1021 cgagatcctc gccgtcgggc atgcgcgcct tgagcctggc gaacagttcg gctggcgcga
     1081 gcccctgatg ctcttcgtcc agatcatcct gatcgacaag accggcttcc atccgagtac
     1141 gtgctcgctc gatgcgatgt ttcgcttggt ggtcgaatgg gcaggtagcc ggatcaagcg
     1201 tatgcagccg ccgcattgca tcagccatga tggatacttt ctcggcagga gcaaggtgag
     1261 atgacaggag atcctgcccc ggcacttcgc ccaatagcag ccagtccctt cccgcttcag
     1321 tgacaacgtc gagcacagct gcgcaaggaa cgcccgtcgt ggccagccac gatagccgcg
     1381 ctgcctcgtc ctgcagttca ttcagggcac cggacaggtc ggtcttgaca aaaagaaccg
     1441 ggcgcccctg cgctgacagc cggaacacgg cggcatcaga gcagccgatt gtctgttgtg
     1501 cccagtcata gccgaatagc ctctccaccc aagcggccgg agaacctgcg tgcaatccat
     1561 cttgttcaat catgcgaaac gatcctcatc ctgtctcttg atcagatccg aaaatggata
     1621 tccaagctcc cgggagcttt ttgcaaaagc ctaggcctcc aaaaaagcct cctcactact
     1681 tctggaatag ctcagaggcc gaggcggcct cggcctctgc ataaataaaa aaaattagtc
     1741 agccatgggg cggagaatgg gcggaactgg gcggagttag gggcgggatg ggcggagtta
     1801 ggggcgggac tatggttgct gactaattga gatgcatgct ttgcatactt ctgcctgctg
     1861 gggagcctgg ggactttcca cacctggttg ctgactaatt gagatgcatg ctttgcatac
     1921 ttctgcctgc tggggagcct ggggactttc cacaccctaa ctgacacaca ttccacagaa
     1981 ttccatcaca aagctctgac ctcaatccta tagaaaggag gaatgagcca aaattcaccc
     2041 aacttattgt gggaagcttg atatccatgg aattcactag tccatagagc ccaccgcatc
     2101 cccagcatgc ctgctattgt cttcccaatc ctcccccttg ctgtcctgcc ccaccccacc
     2161 ccccagaata gaatgacacc tactcagaca atgcgatgca atttcctcat tttattagga
     2221 aaggacagtg ggagtggcac cttccagggt caaggaaggc acgggggagg ggcaaacaac
     2281 agatggctgg caactagaag gcacagtcga ggctgatcag cgggtttaaa cttaggaccc
     2341 actttcacat ttaagttgtt tttctaatcc gcatatgatc aattcaaggc cgaataagaa
     2401 ggctggctct gcaccttggt gatcaaataa ttcgatagct tgtcgtaata atggcggcat
     2461 actatcagta gtaggtgttt ccctttcttc tttagcgact tgatgctctt gatcttccaa
     2521 tacgcaacct aaagtaaaat gccccacagc gctgagtgca tataatgcat tctctagtga
     2581 aaaaccttgt tggcataaaa aggctaattg attttcgaga gtttcatact gtttttctgt
     2641 aggccgtgta cctaaatgta cttttgctcc atcgcgatga cttagtaaag cacatctaaa
     2701 acttttagcg ttattacgta aaaaatcttg ccagctttcc ccttctaaag ggcaaaagtg
     2761 agtatggtgc ctatctaaca tctcaatggc taaggcgtcg agcaaagccc gcttattttt
     2821 tacatgccaa tacaatgtag gctgctctac acctagcttc tgggcgagtt tacgggttgt
     2881 taaaccttcg attccgacct cattaagcag ctctaatgcg ctgttaatca ctttactttt
     2941 atctaatcta gacatatcaa ttctttgcca aaatgatgag acagcacaat aaccagcacg
     3001 ttgcggccgg agctgtagga aaaagaagaa ggcatgaaca tggttagcag aggggcccgg
     3061 tttggactca gagtatttta tcctcatctc aaacagtgta tatcattgta accataaaga
     3121 gaaaggcagg atgatgacca ggatgtagtt gtttctacca ataagaatat ttccacgcca
     3181 gccagaattt atatgcagaa atattctacc ttatcattta attataacaa ttgttctcta
     3241 aaactgtgct gaagtacaat ataatatacc ctgattgcct tgaaaaaaaa gtgattagag
     3301 aaagtactta caatctgaca aataaacaaa agtgaattta aaaattcgtt acaaatgcaa
     3361 gctaaagttt aacgaaaaag ttacagaaaa tgaaaagaaa ataagaggag acaatggttg
     3421 tcaacagagt agaaagtgaa agaaacaaaa ttatcatgag ggtccatggt gatacaaggg
     3481 acatcttccc attctaaaca acaccctgaa aactttgccc cctccatata acatgaattt
     3541 tacaatagcg aaaaagaaag aacaatcaag ggtccccaaa ctcaccctga agttctcagg
     3601 atcgctcgag ctttttgcaa aagcctaggc ctccaaaaaa gcctcctcac tacttctgga
     3661 atagctcaga ggccgaggcg gcctcggcct ctgcataaat aaaaaaaatt agtcagccat
     3721 ggggcggaga atgggcggaa ctgggcggag ttaggggcgg gatgggcgga gttaggggcg
     3781 ggactatggt tgctgactaa ttgagatgca tgctttgcat acttctgcct gctggggagc
     3841 ctggggactt tccacacctg gttgctgact aattgagatg catgctttgc atacttctgc
     3901 ctgctgggga gcctggggac tttccacacc ctaactgaca cacattccac agactagtgc
     3961 gcgcggccgc tctagatggc cagatctagc ttgtggaagg ctactcgaaa tgtttgaccc
     4021 aagttaaaca atttaaaggc aatgctacca aatactaatt gagtgtatgt aaacttctga
     4081 cccactggga atgtgatgaa agaaataaaa gctgaaatga atcattctct ctactattat
     4141 tctgatattt cacattctta aaataaagtg gtgatcctaa ctgacctaag acagggaatt
     4201 tttactagga ttaaatgtca ggaattgtga aaaagtgagt ttaaatgtat ttggctaagg
     4261 tgtatgtaaa cttccgactt caactgtata gggatcctct agctagagtc gacctcgagg
     4321 gggggcccgg tacccagctt ttgttccctt tagtgagggt taattgcgcg cttggcgtaa
     4381 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     4441 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     4501 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     4561 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     4621 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     4681 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     4741 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     4801 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     4861 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     4921 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaaccgt ggcgctttct
     4981 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     5041 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     5101 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     5161 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     5221 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     5281 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     5341 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     5401 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     5461 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     5521 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     5581 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     5641 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     5701 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     5761 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     5821 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     5881 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     5941 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     6001 agtaacttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     6061 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     6121 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     6181 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     6241 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     6301 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     6361 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     6421 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     6481 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     6541 gcgccctgta gcggcgcatt aagcgcggcg ggagtggtgg ttacgcgcag cgtgaccgct
     6601 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     6661 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     6721 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     6781 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     6841 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     6901 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     6961 gcgaatttta acaaaatatt aacgcttaca atttccattc gccattcagg ctgcgcaact
     7021 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
     7081 gtgctg
//
