LOCUS       Exported                7706 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  expresses two human WT p53s tethered together and also expresses 
            EGFP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pIRES2-EGFP-p53 tethered
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7706)
  AUTHORS   Lin SC, Karoly ED, Taatjes DJ
  TITLE     The human DeltaNp53 isoform triggers metabolic and gene expression 
            changes that activate mTOR and alter mitochondrial function.
  JOURNAL   Aging Cell. 2013 Oct;12(5):863-72. doi: 10.1111/acel.12108. Epub 
            2013 Jun 25.
  PUBMED    23734707
REFERENCE   2  (bases 1 to 7706)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7706
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1838..1858
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1868..3052
                     /codon_start=1
                     /gene="Homo sapiens TP53"
                     /product="human tumor suppressor protein that is often 
                     mutated in cancers"
                     /label=p53
                     /note="responds to cellular stresses to regulate the 
                     expression of target genes"
                     /translation="MEEPQSDPSVEPPLSQETFSDLWKLLPENNVLSPLPSQAMDDLML
                     SPDDIEQWFTEDPGPDEAPRMPEAAPPVAPAPAAPTPAAPAPAPSWPLSSSVPSQKTYQ
                     GSYGFRLGFLHSGTAKSVTCTYSPALNKMFCQLAKTCPVQLWVDSTPPPGTRVRAMAIY
                     KQSQHMTEVVRRCPHHERCSDSDGLAPPQHLIRVEGNLRVEYLDDRNTFRHSVVVPYEP
                     PEVGSDCTTIHYNYMCNSSCMGGMNRRPILTIITLEDSSGNLLGRNSFEVRVCACPGRD
                     RRTEEENLRKKGEPHHELPPGSTKRALPNNTSSSPQPKKKPLDGEYFTLQIRGRERFEM
                     FRELNEALELKDAQAGKEPGGSRAHSSHLKSKKGQSTSRHKKLMFKTEGPDSDG"
     misc_feature    3065..3651
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3232..3249)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3459..3478
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3652..4371
                     /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             3652..4368
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3697..3718)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3958..3977)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4305..4326
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4494..4615
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4531..4550)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4585..4604
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4622..5077)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4759..4780)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4971..4990
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5104..5208
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5210..5567
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(5233..5253)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5418..5553
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5480..5499
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5602..6396
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5656..5675)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6266..6285
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6584..6603)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6628..6675
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7004..7592
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7493..7512
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttcga attccacctg ctaggtatgg aagagccgca
      661 gtcagatcct agcgtcgagc cccctctgag tcaggaaaca ttttcagacc tatggaaact
      721 acttcctgaa aacaacgttc tgtccccctt gccgtcccaa gcaatggatg atttgatgct
      781 gtccccggac gatattgaac aatggttcac tgaagaccca ggtccagatg aagctcccag
      841 aatgccagag gctgctcccc ccgtggcccc tgcaccagca gctcctacac cggcggcccc
      901 tgcaccagcc ccctcctggc ccctgtcatc ttctgtccct tcccagaaaa cctaccaggg
      961 cagctacggt ttccgtctgg gcttcttgca ttctgggaca gccaagtctg tgacttgcac
     1021 gtactcccct gccctcaaca agatgttttg ccaactggcc aagacctgcc ctgtgcagct
     1081 gtgggttgat tccacacccc cgcccggcac ccgcgtccgc gccatggcca tctacaagca
     1141 gtcacagcac atgacggagg ttgtgaggcg ctgcccccac catgagcgct gctcagatag
     1201 cgatggtctg gcccctcctc agcatcttat ccgagtggaa ggaaatttgc gtgtggagta
     1261 tttggatgac agaaacactt ttcgacatag tgtggtggtg ccctatgagc cgcctgaggt
     1321 tggctctgac tgtaccacca tccactacaa ctacatgtgt aacagttcct gcatgggcgg
     1381 catgaaccgg aggcccatcc tcaccatcat cacactggaa gactccagtg gtaatctact
     1441 gggacggaac agctttgagg tgcgtgtttg tgcctgtcct gggagagacc ggcgcacaga
     1501 ggaagagaat ctccgcaaga aaggggagcc tcaccacgag ctgcccccag ggagcactaa
     1561 gcgagcactg cccaacaaca ccagctcctc tccccagcca aagaagaaac cactggatgg
     1621 agaatatttc acccttcaga tccgtgggcg tgagcgcttc gagatgttcc gagagctgaa
     1681 tgaggccttg gaactcaagg atgcccaggc tgggaaggag ccagggggga gcagggctca
     1741 ctccagccac ctgaagtcca aaaagggtca gtctacctcc cgccataaaa aactcatgtt
     1801 caagacagaa gggccagact ctgacggtgg cggaggggaa aatctgtatt tccagggagg
     1861 cggaggtatg gaagagccgc agtcagatcc tagcgtcgag ccccctctga gtcaggaaac
     1921 attttcagac ctatggaaac tacttcctga aaacaacgtt ctgtccccct tgccgtccca
     1981 agcaatggat gatttgatgc tgtccccgga cgatattgaa caatggttca ctgaagaccc
     2041 aggtccagat gaagctccca gaatgccaga ggctgctccc cccgtggccc ctgcaccagc
     2101 agctcctaca ccggcggccc ctgcaccagc cccctcctgg cccctgtcat cttctgtccc
     2161 ttcccagaaa acctaccagg gcagctacgg tttccgtctg ggcttcttgc attctgggac
     2221 agccaagtct gtgacttgca cgtactcccc tgccctcaac aagatgtttt gccaactggc
     2281 caagacctgc cctgtgcagc tgtgggttga ttccacaccc ccgcccggca cccgcgtccg
     2341 cgccatggcc atctacaagc agtcacagca catgacggag gttgtgaggc gctgccccca
     2401 ccatgagcgc tgctcagata gcgatggtct ggcccctcct cagcatctta tccgagtgga
     2461 aggaaatttg cgtgtggagt atttggatga cagaaacact tttcgacata gtgtggtggt
     2521 gccctatgag ccgcctgagg ttggctctga ctgtaccacc atccactaca actacatgtg
     2581 taacagttcc tgcatgggcg gcatgaaccg gaggcccatc ctcaccatca tcacactgga
     2641 agactccagt ggtaatctac tgggacggaa cagctttgag gtgcgtgttt gtgcctgtcc
     2701 tgggagagac cggcgcacag aggaagagaa tctccgcaag aaaggggagc ctcaccacga
     2761 gctgccccca gggagcacta agcgagcact gcccaacaac accagctcct ctccccagcc
     2821 aaagaagaaa ccactggatg gagaatattt cacccttcag atccgtgggc gtgagcgctt
     2881 cgagatgttc cgagagctga atgaggcctt ggaactcaag gatgcccagg ctgggaagga
     2941 gccagggggg agcagggctc actccagcca cctgaagtcc aaaaagggtc agtctacctc
     3001 ccgccataaa aaactcatgt tcaagacaga agggcccgac tcagacgggt agatatcgga
     3061 tccgcccctc tccctccccc ccccctaacg ttactggccg aagccgcttg gaataaggcc
     3121 ggtgtgcgtt tgtctatatg ttattttcca ccatattgcc gtcttttggc aatgtgaggg
     3181 cccggaaacc tggccctgtc ttcttgacga gcattcctag gggtctttcc cctctcgcca
     3241 aaggaatgca aggtctgttg aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa
     3301 gacaaacaac gtctgtagcg accctttgca ggcagcggaa ccccccacct ggcgacaggt
     3361 gcctctgcgg ccaaaagcca cgtgtataag atacacctgc aaaggcggca caaccccagt
     3421 gccacgttgt gagttggata gttgtggaaa gagtcaaatg gctctcctca agcgtattca
     3481 acaaggggct gaaggatgcc cagaaggtac cccattgtat gggatctgat ctggggcctc
     3541 ggtgcacatg ctttacatgt gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc
     3601 acggggacgt ggttttcctt tgaaaaacac gatgataata tggccacaac catggtgagc
     3661 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta
     3721 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
     3781 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
     3841 accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac
     3901 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
     3961 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
     4021 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
     4081 tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag
     4141 gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac
     4201 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
     4261 acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
     4321 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta aagcggccgc
     4381 gactctagat cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc
     4441 tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt
     4501 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     4561 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttaaggcg
     4621 taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca
     4681 ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag
     4741 atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc
     4801 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc
     4861 taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc
     4921 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa
     4981 gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc
     5041 acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat
     5101 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     5161 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa
     5221 agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     5281 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     5341 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     5401 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     5461 atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat
     5521 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaagat cgatcaagag
     5581 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     5641 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     5701 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     5761 tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg
     5821 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     5881 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     5941 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     6001 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     6061 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     6121 ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     6181 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     6241 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     6301 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     6361 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga
     6421 ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg
     6481 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     6541 atctcatgct ggagttcttc gcccacccta gggggaggct aactgaaaca cggaaggaga
     6601 caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacggtg
     6661 ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc
     6721 caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc
     6781 caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag
     6841 cctcaggtta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     6901 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     6961 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     7021 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     7081 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     7141 caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     7201 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     7261 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     7321 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     7381 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     7441 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     7501 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     7561 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     7621 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     7681 tggataaccg tattaccgcc atgcat
//
