LOCUS       sequence-198999-       10225 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10225)
  AUTHORS   Yu J, Zhang L, Hwang PM, Kinzler KW, Vogelstein B
  TITLE     PUMA induces the rapid apoptosis of colorectal cancer cells.
  JOURNAL   Mol Cell. 2001 Mar . 7(3):673-82.
  PUBMED    11463391
REFERENCE   2  (bases 1 to 10225)
  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: "Mol Cell. 
            2001 Mar . 7(3):673-82."
COMMENT     Sequence Label: pHA-PUMA
FEATURES             Location/Qualifiers
     source          1..10225
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(781..799)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        867..971
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(972..1040,1041..1832)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1190..1209)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2003..2591
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2492..2511
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3022..3167
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     CDS             3210..4247
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPDRE
                     MGEAN"
     primer_bind     complement(4245..4264)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4289..4337
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     enhancer        4991..5370
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5371..5574
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5525..5545
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5571..5595
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             5646..5672
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     polyA_signal    6320..6454
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6344..6363)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6398..6417
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(6597..6619)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     rep_origin      7179..8968
                     /label=oriP
                     /note="Epstein-Barr virus oriP replication origin (Yates et
                     al., 2000)"
ORIGIN
        1 ctcctgcccc tcctgctcct gcccctcctc ctgctcctgc ccctcctgct cctgcccctc
       61 ccgctcctgc tcctgctcct gttccaccgt gggtcccttt gcagccaatg caacttggac
      121 gtttttgggg tctccggaca ccatctctat gtcttggccc tgatcctgag ccgcccgggg
      181 ctcctggtct tccgcctcct cgtcctcgtc ctcttccccg tcctcgtcca tggttatcac
      241 cccctcttct ttgaggtcca ctgccgccgg agccttctgg tccagatgtg tctcccttct
      301 ctcctaggcc atttccaggt cctgtacctg gcccctcgtc agacatgatt cacactaaaa
      361 gagatcaata gacatcttta ttagacgacg ctcagtgaat acagggagtg cagactcctg
      421 ccccctccaa cagccccccc accctcatcc ccttcatggt cgctgtcaga cagatccagg
      481 tctgaaaatt ccccatcctc cgaaccatcc tcgtcctcat caccaattac tcgcagcccg
      541 gaaaactccc gctgaacatc ctcaagattt gcgtcctgag cctcaagcca ggcctcaaat
      601 tcctcgtccc cctttttgct ggacggtagg gatggggatt ctcgggaccc ctcctcttcc
      661 tcttcaaggt caccagacag agatgctact ggggcaacgg aagaaaagct gggtgcggcc
      721 tgtgaggatc agcttatcga tgataagctg tcaaacatga gaattcttga agacgaaagg
      781 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
      841 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
      901 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
      961 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     1021 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     1081 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     1141 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     1201 cggtattatc ccgtgttgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     1261 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     1321 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     1381 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     1441 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     1501 acaccacgat gcctgcagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     1561 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     1621 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     1681 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     1741 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     1801 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     1861 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     1921 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     1981 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     2041 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     2101 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     2161 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     2221 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     2281 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     2341 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     2401 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     2461 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     2521 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     2581 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     2641 gaagctgtcc ctgatggtcg tcatctacct gcctggacag catggcctgc aacgcgggca
     2701 tcccgatgcc gccggaagcg agaagaatca taatggggaa ggccatccag cctcgcgtcg
     2761 cgaacgccag caagacgtag cccagcgcgt cggccccgag atgcgccgcg tgcggctgct
     2821 ggagatggcg gacgcgatgg atatgttctg ccaagggttg gtttgcgcat tcacagttct
     2881 ccgcaagaat tgattggctc caattcttgg agtggtgaat ccgttagcga ggtgccgccc
     2941 tgcttcatcc ccgtggcccg ttgctcgcgt ttgctggcgg tgtccccgga agaaatatat
     3001 ttgcatgtct ttagttctat gatgacacaa accccgccca gcgtcttgtc attggcgaat
     3061 tcgaacacgc agatgcagtc ggggcggcgc ggtccgaggt ccacttcgca tattaaggtg
     3121 acgcgtgtgg cctcgaacac cgagcgaccc tgcagcgacc cgcttaacag cgtcaacagc
     3181 gtgccgcaga tcccgggggg caatgagata tgaaaaagcc tgaactcacc gcgacgtctg
     3241 tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga cctgatgcag ctctcggagg
     3301 gcgaagaatc tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa
     3361 atagctgcgc cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatcggccg
     3421 cgctcccgat tccggaagtg cttgacattg gggaattcag cgagagcctg acctattgca
     3481 tctcccgccg tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg
     3541 ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc ggccgatctt agccagacga
     3601 gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca
     3661 tatgcgcgat tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca
     3721 gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag
     3781 tccggcacct cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca
     3841 taacagcggt cattgactgg agcgaggcga tgttcgggga ttcccaatac gaggtcgcca
     3901 acatcttctt ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc
     3961 ggaggcatcc ggagcttgca ggatcgccgc ggctccgggc gtatatgctc cgcattggtc
     4021 ttgaccaact ctatcagagc ttggttgacg gcaatttcga tgatgcagct tgggcgcagg
     4081 gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca caaatcgccc
     4141 gcagaagcgc ggccgtctgg accgatggct gtgtagaagt actcgccgat agtggaaacc
     4201 gacgccccag cactcgtccg gatcgggaga tgggggaggc taactgaaac acggaaggag
     4261 acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggg
     4321 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac
     4381 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc
     4441 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat
     4501 agccactggc cccgtgggtt agggacgggg tcccccatgg ggaatggttt atggttcgtg
     4561 ggggttatta ttttgggcgt tgcgtggggt caggtccacg actggactga gcagacagac
     4621 ccatggtttt tggatggcct gggcatggac cgcatgtact ggcgcgacac gaacaccggg
     4681 cgtctgtggc tgccaaacac ccccgacccc caaaaaccac cgcgcggatt tctggcgtgc
     4741 caagctagtc gaccaattct catgtttgac agcttatcat cgcagatccg ggcaacgttg
     4801 ttgccattgc tgcaggcgca gaactggtag gtatggaaga tctatacatt gaatcaatat
     4861 tggcaattag ccatattagt cattggttat atagcataaa tcaatattgg ctattggcca
     4921 ttgcatacgt tgtatctata tcataatatg tacatttata ttggctcatg tccaatatga
     4981 ccgccatgtt gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
     5041 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
     5101 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
     5161 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     5221 gcagtacatc aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa
     5281 tggcccgcct ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac
     5341 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg
     5401 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     5461 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg
     5521 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta
     5581 gtgaaccgtc agatctctag aagctgggta ccgaaatggg atacgatgtt ccagattacg
     5641 cttcgtacga tgttccagat tacgcaagct tgatggcccg cgcacgccag gagggcagct
     5701 ccccggagcc cgtagagggc ctggcccgcg acggcccgcg ccccttcccg ctcggccgcc
     5761 tggtgccctc ggcagtgtcc tgcggcctct gcgagcccgg cctggctgcc gcccccgccg
     5821 cccccaccct gctgcccgct gcctacctct gcgcccccac cgccccaccc gccgtcaccg
     5881 ccgccctggg gggttcccgc tggcctgggg gtccccgcag ccggccccga ggcccgcgcc
     5941 cggacggtcc tcagccctcg ctctcgctgg cggagcagca cctggagtcg cccgtgccca
     6001 gcgccccggg ggctctggcg ggcggtccca cccaggcggc cccgggagtc cgcggggagg
     6061 aggaacagtg ggcccgggag atcggggccc agctgcggcg gatggcggac gacctcaacg
     6121 cacagtacga gcggcggaga caagaggagc agcagcggca ccgcccctca ccctggaggg
     6181 tcctgtacaa tctcatcatg ggactcctgc ccttacccag gggccacaga gcccccgaga
     6241 tggagcccaa ttaggtgcct gcacccgccc ggtggacgtc agggactcgg ggggcaggcc
     6301 cctcccacct cctgataagg atccagacat gataagatac attgatgagt ttggacaaac
     6361 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt
     6421 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat
     6481 gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg
     6541 tggtatggct gattatgatc cggctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc
     6601 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
     6661 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc catgaggtcg
     6721 actctagagg atcgatcccc gccccggacg aactaaacct gactacgaca tctctgcccc
     6781 ttcttcgcgg ggcagtgcat gtaatccctt cagttggttg gtacaacttg ccaactgggc
     6841 cctgttccac atgtgacacg gggggggacc aaacacaaag gggttctctg actgtagttg
     6901 acatccttat aaatggatgt gcacatttgc caacactgag tggctttcat cctggagcag
     6961 actttgcagt ctgtggactg caacacaaca ttgcctttat gtgtaactct tggctgaagc
     7021 tcttacacca atgctggggg acatgtacct cccaggggcc caggaagact acgggaggct
     7081 acaccaacgt caatcagagg ggcctgtgta gctaccgata agcggaccct caagagggca
     7141 ttagcaatag tgtttataag gcccccttgt taaccctaaa cgggtagcat atgcttcccg
     7201 ggtagtagta tatactatcc agactaaccc taattcaata gcatatgtta cccaacggga
     7261 agcatatgct atcgaattag ggttagtaaa agggtcctaa ggaacagcga tatctcccac
     7321 cccatgagct gtcacggttt tatttacatg gggtcaggat tccacgaggg tagtgaacca
     7381 ttttagtcac aagggcagtg gctgaagatc aaggagcggg cagtgaactc tcctgaatct
     7441 tcgcctgctt cttcattctc cttcgtttag ctaatagaat aactgctgag ttgtgaacag
     7501 taaggtgtat gtgaggtgct cgaaaacaag gtttcaggtg acgcccccag aataaaattt
     7561 ggacgggggg ttcagtggtg gcattgtgct atgacaccaa tataaccctc acaaacccct
     7621 tgggcaataa atactagtgt aggaatgaaa cattctgaat atctttaaca atagaaatcc
     7681 atggggtggg gacaagccgt aaagactgga tgtccatctc acacgaattt atggctatgg
     7741 gcaacacata atcctagtgc aatatgatac tggggttatt aagatgtgtc ccaggcaggg
     7801 accaagacag gtgaaccatg ttgttacact ctatttgtaa caaggggaaa gagagtggac
     7861 gccgacagca gcggactcca ctggttgtct ctaacacccc cgaaaattaa acggggctcc
     7921 acgccaatgg ggcccataaa caaagacaag tggccactct tttttttgaa attgtggagt
     7981 gggggcacgc gtcagccccc acacgccgcc ctgcggtttt ggactgtaaa ataagggtgt
     8041 aataacttgg ctgattgtaa ccccgctaac cactgcggtc aaaccacttg cccacaaaac
     8101 cactaatggc accccgggga atacctgcat aagtaggtgg gcgggccaag ataggggcgc
     8161 gattgctgcg atctggagga caaattacac acacttgcgc ctgagcgcca agcacagggt
     8221 tgttggtcct catattcacg aggtcgctga gagcacggtg ggctaatgtt gccatgggta
     8281 gcatatacta cccaaatatc tggatagcat atgctatcct aatctatatc tgggtagcat
     8341 aggctatcct aatctatatc tgggtagcat atgctatcct aatctatatc tgggtagtat
     8401 atgctatcct aatttatatc tgggtagcat aggctatcct aatctatatc tgggtagcat
     8461 atgctatcct aatctatatc tgggtagtat atgctatcct aatctgtatc cgggtagcat
     8521 atgctatcct aatagagatt agggtagtat atgctatcct aatttatatc tgggtagcat
     8581 atactaccca aatatctgga tagcatatgc tatcctaatc tatatctggg tagcatatgc
     8641 tatcctaatc tatatctggg tagcataggc tatcctaatc tatatctggg tagcatatgc
     8701 tatcctaatc tatatctggg tagtatatgc tatcctaatt tatatctggg tagcataggc
     8761 tatcctaatc tatatctggg tagcatatgc tatcctaatc tatatctggg tagtatatgc
     8821 tatcctaatc tgtatccggg tagcatatgc tatcctcatg catatacagt cagcatatga
     8881 tacccagtag tagagtggga gtgctatcct ttgcatatgc cgccacctcc caagggggcg
     8941 tgaattttcg ctgcttgtcc ttttcctgct ggttgctccc attcttaggt gaatttaagg
     9001 aggccaggct aaagccgtcg catgtctgat tgctcaccag gtaaatgtcg ctaatgtttt
     9061 ccaacgcgag aaggtgttga gcgcggagct gagtgacgtg acaacatggg tatgcccaat
     9121 tgccccatgt tgggaggacg aaaatggtga caagacagat ggccagaaat acaccaacag
     9181 cacgcatgat gtctactggg gatttattct ttagtgcggg ggaatacacg gcttttaata
     9241 cgattgaggg cgtctcctaa caagttacat cactcctgcc cttcctcacc ctcatctcca
     9301 tcacctcctt catctccgtc atctccgtca tcaccctccg cggcagcccc ttccaccata
     9361 ggtggaaacc agggaggcaa atctactcca tcgtcaaagc tgcacacagt caccctgata
     9421 ttgcaggtag gagcgggctt tgtcataaca aggtccttaa tcgcatcctt caaaacctca
     9481 gcaaatatat gagtttgtaa aaagaccatg aaataacaga caatggactc ccttagcggg
     9541 ccaggttgtg ggccgggtcc aggggccatt ccaaagggga gacgactcaa tggtgtaaga
     9601 cgacattgtg gaatagcaag ggcagttcct cgccttaggt tgtaaaggga ggtcttacta
     9661 cctccatata cgaacacacc ggcgacccaa gttccttcgt cggtagtcct ttctacgtga
     9721 ctcctagcca ggagagctct taaaccttct gcaatgttct caaatttcgg gttggaacct
     9781 ccttgaccac gatgctttcc aaaccaccct ccttttttgc gcctgcctcc atcaccctga
     9841 ccccggggtc cagtgcttgg gccttctcct gggtcatctg cggggccctg ctctatcgct
     9901 cccgggggca cgtcaggctc accatctggg ccaccttctt ggtggtattc aaaataatcg
     9961 gcttccccta cagggtggaa aaatggcctt ctacctggag ggggcctgcg cggtggagac
    10021 ccggatgatg atgactgact actgggactc ctgggcctct tttctccacg tccacgacct
    10081 ctccccctgg ctctttcacg acttcccccc ctggctcttt cacgtcctct accccggcgg
    10141 cctccactac ctcctcgacc ccggcctcca ctacctcctc gaccccggcc tccactgcct
    10201 cctcgacccc ggcctccacc tcctg
//
