LOCUS       sequence-343323-        7613 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TORCAR targeted to the nucleus via histone H2A..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7613)
  AUTHORS   Zhou X, Zhong Y, Molinar-Inglis O, Kunkel MT, Chen M, Sun T, Zhang 
            J, Shyy JY, Trejo J, Newton AC, Zhang J
  TITLE     Location-specific inhibition of Akt reveals regulation of mTORC1 
            activity in the nucleus.
  JOURNAL   Nat Commun. 2020 Nov 30;11(1):6088. doi: 10.1038/s41467-020-19937-w.
  PUBMED    33257668
REFERENCE   2  (bases 1 to 7613)
  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: "Nat 
            Commun."; date: "2020-11-30"; volume: "11(1):6088. doi"; pages: " 
            10.1038/s41467-020-19937-w"
COMMENT     Sequence Label: pcDNA3-H2A-TORCAR
FEATURES             Location/Qualifiers
     source          1..7613
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(23..42)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        214..593
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        594..797
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     748..768
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     842..861
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        842..860
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(1282..1284,1285..1287,1288..1998)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     AHFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1327..1348)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1588..1607)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1935..1956
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2368..3078
                     /codon_start=1
                     /product="yellow fluorescent protein optimized for FRET 
                     with CyPet (Nguyen and Daugherty, 2005)"
                     /label=YPet
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKL
                     LCTTGKLPVPWPTLVTTLGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKAN
                     FKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALFKDPNEKRDHMVLLEF
                     LTAAGITEGMNELYK"
     promoter        complement(3145..3163)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3146..3163)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3183..3200)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3189..3413
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3459..3887
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3546..3565)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3756..3777
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3896..3916)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3901..4230
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4081..4216
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4143..4162
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4297..5091
                     /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(4351..4370)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4961..4980
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5265..5386
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5302..5321)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5356..5375
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5435..5451)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5435..5451)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5448..5470)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5459..5475
                     /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(5483..5489,5490..5507,5508..5513))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5528..5549
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5666..5683)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5837..6425)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5917..5936)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6596..7387,7388..7456))
                     /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     7219..7238
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7457..7561)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc
       61 tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa
      121 caaggcaagg cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc
      181 tgcttcgcga tgtacgggcc agatatacgc gttgacattg attattgact agttattaat
      241 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
      301 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
      361 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      421 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
      481 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
      541 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
      601 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
      661 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
      721 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg
      781 tctatataag cagagctctc tggctaacta gagaacccac tgcttactgg cttatcgaaa
      841 ttaatacgac tcactatagg gagacccaag cttgcggccg ccaccatgtc tggtcgtggc
      901 aaacaaggag gcaaggcccg cgccaaggcc aagtcgcgct cgtcccgcgc tggcctccag
      961 ttcccggtag ggcgagtgca ccgcttgctg cgcaaaggca actacgcgga gcgggtgggg
     1021 gccggcgcgc ccgtctacat ggcggcggtc ctcgagtacc tgaccgccga gatcctggag
     1081 ctggcgggca acgcggctcg ggacaacaag aagacgcgca tcatccctcg tcacctccag
     1141 ctggccatcc gcaacgacga ggaactgaac aagctgctgg gcaaagtcac catcgcccag
     1201 ggcggcgttt tgcctaacat ccaggccgtt ctgttaccaa agaaaaccga aagccacaaa
     1261 gccaaaagca aagcggatcc catggtgagc aagggcgagg agctgttcac cggggtggtg
     1321 cccatcctgg tcgagctgga cggcgacgta aacggccaca ggttcagcgt gtccggcgag
     1381 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
     1441 ctgcccgtgc cctggcccac cctcgtgacc accctgacct ggggcgtgca gtgcttcgcc
     1501 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     1561 gtccaggagc gtaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     1621 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     1681 gacggcaaca tcctggggca caagctggag tacaacgcca tcagtgacaa cgtctatatc
     1741 accgccgaca agcagaagaa cggcatcaag gcccacttca agatccgcca caacatcgag
     1801 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     1861 gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac
     1921 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     1981 atggacgagc tgtacaagcg catgcatatg tccgggggca gcagctgcag ccagacccca
     2041 agccgggcca tccccgccac tcgccgggtg gtgctcggcg acggcgtgca gctcccgccc
     2101 ggggactaca gcacgacccc cggcggcacg ctcttcagca ccaccccggg aggtaccagg
     2161 atcatctatg accggaaatt cctgatggag tgtcggaact cacctgtgac caaaacaccc
     2221 ccaagggatc tgcccaccat tccgggggtc accagccctt ccagtgatga gccccccatg
     2281 gaagccagcc agagccacct gcgcaatagc ccagaagata agcgggcggg cggtgaagag
     2341 tcacagtttg agatggacat tgagctctct aaaggtgaag aattattcac tggtgttgtc
     2401 ccaattttgg ttgaattaga tggtgatgtt aatggtcaca aattttctgt ctccggtgaa
     2461 ggtgaaggtg atgctacgta cggtaaattg accttaaaat tactctgtac tactggtaaa
     2521 ttgccagttc catggccaac cttagtcact actttaggtt atggtgttca atgttttgct
     2581 agatacccag atcatatgaa acaacatgac tttttcaagt ctgccatgcc agaaggttat
     2641 gttcaagaaa gaactatttt tttcaaagat gacggtaact acaagaccag agctgaagtc
     2701 aagtttgaag gtgatacctt agttaataga atcgaattaa aaggtattga ttttaaagaa
     2761 gatggtaaca ttttaggtca caaattggaa tacaactata actctcacaa tgtttacatc
     2821 actgctgaca aacaaaagaa tggtatcaaa gctaacttca aaattagaca caacattgaa
     2881 gatggtggtg ttcaattagc tgaccattat caacaaaata ctccaattgg tgatggtcca
     2941 gtcttgttac cagacaacca ttacttatcc tatcaatctg ccttattcaa agatccaaac
     3001 gaaaagagag accacatggt cttgttagaa tttttgactg ctgctggtat taccgagggt
     3061 atgaatgaat tgtacaaata gtaagaattc tgcagatatc catcacactg gcggccgctc
     3121 gagcatgcat ctagagggcc ctattctata gtgtcaccta aatgctagag ctcgctgatc
     3181 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     3241 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     3301 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     3361 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     3421 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     3481 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3541 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3601 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3661 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3721 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     3781 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     3841 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     3901 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3961 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     4021 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     4081 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     4141 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     4201 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     4261 ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     4321 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     4381 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     4441 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
     4501 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     4561 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     4621 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     4681 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     4741 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     4801 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat
     4861 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     4921 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     4981 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     5041 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     5101 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     5161 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     5221 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag
     5281 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     5341 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac
     5401 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     5461 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     5521 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     5581 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     5641 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     5701 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     5761 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     5821 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     5881 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     5941 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     6001 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     6061 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     6121 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     6181 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     6241 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     6301 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     6361 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     6421 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     6481 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     6541 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     6601 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     6661 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     6721 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     6781 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     6841 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     6901 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     6961 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     7021 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     7081 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     7141 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     7201 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     7261 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     7321 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     7381 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     7441 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     7501 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     7561 gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc ccg
//
