LOCUS       sequence-214277-        8485 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of p14ARF IRES H2B-mOrange2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8485)
  AUTHORS   Zeng H, Jorapur A, Shain AH, Lang UE, Torres R, Zhang Y, McNeal AS, 
            Botton T, Lin J, Donne M, Bastian IN, Yu R, North JP, Pincus L, 
            Ruben BS, Joseph NM, Yeh I, Bastian BC, Judson RL
  TITLE     Bi-allelic Loss of CDKN2A Initiates Melanoma Invasion via BRN2 
            Activation.
  JOURNAL   Cancer Cell. 2018 Jul 9;34(1):56-68.e9. doi: 
            10.1016/j.ccell.2018.05.014.
  PUBMED    29990501
REFERENCE   2  (bases 1 to 8485)
  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"; doi: 
            "10.1016/j.ccell.2018.05"; journalName: "Cancer Cell"; date: 
            "2018-07-9- 9"; volume: "34"; issue: "1"; pages: "56-68.e9"
COMMENT     Sequence Label: pHIV-ARF-mOrange2
FEATURES             Location/Qualifiers
     source          1..8485
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(50..69)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        241..620
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        622..820
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     775..795
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             838..1018
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1065..1190
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1687..1920
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2105..2149
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2447..2564
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2627..3805
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2858..3796
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3753..3773
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     misc_feature    4265..4838
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    4288..4840
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4432..4449)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4659..4678
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4844..5221
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     regulatory      5234..5243
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5240..5950
                     /codon_start=1
                     /product="photostable monomeric orange derivative of DsRed 
                     fluorescent protein (Shaner et al., 2008)"
                     /label=mOrange2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG
                     FQTAKLKVTKGGPLPFAWDILSPHFTYGSKAYVKHPADIPDYFKLSFPEGFKWERVMNY
                     GDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GKIKMRLKLKDGGHYTSEVKTTYKAKKPVQLPGAYIVDIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(5968..5984)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5969..5985)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             6494..6674
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6736..7324)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6816..6835)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7495..8286,8287..8355))
                     /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     8118..8137
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8356..8460)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt acaatctgct
       61 ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt
      121 agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga
      181 atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt
      241 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
      301 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      361 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      421 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      481 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      541 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      601 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      661 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      721 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      781 tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg
      841 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
      901 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
      961 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
     1021 ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga
     1081 ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa
     1141 aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc
     1201 gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat
     1261 ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg
     1321 gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc
     1381 agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc
     1441 atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa
     1501 acaaaagtaa gaccaccgca cagcaagcgg ccggccgctg atcttcagac ctggaggagg
     1561 agatatgagg gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc
     1621 attaggagta gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt
     1681 gggaatagga gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc
     1741 gtcaatgacg ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa
     1801 caatttgctg agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat
     1861 caagcagctc caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct
     1921 ggggatttgg ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag
     1981 ttggagtaat aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag
     2041 agaaattaac aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca
     2101 agaaaagaat gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt
     2161 taacataaca aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt
     2221 aggtttaaga atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc
     2281 accattatcg tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat
     2341 agaagaagaa ggtggagaga gagacagaga cagatccatt cgattagtga acggatcggc
     2401 actgcgtgcg ccaattctgc agacaaatgg cagtattcat ccacaatttt aaaagaaaag
     2461 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2521 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
     2581 acagcagaga tccagtttgg ttagtaccgg gcccgctcta gcgtgaggct ccggtgcccg
     2641 tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa
     2701 ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg
     2761 gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa
     2821 cgttcttttt cgcaacgggt ttgccgccag aacacaggta agtgccgtgt gtggttcccg
     2881 cgggcctggc ctctttacgg gttatggccc ttgcgtgcct tgaattactt ccacctggct
     2941 gcagtacgtg attcttgatc ccgagcttcg ggttggaagt gggtgggaga gttcgaggcc
     3001 ttgcgcttaa ggagcccctt cgcctcgtgc ttgagttgag gcctggcctg ggcgctgggg
     3061 ccgccgcgtg cgaatctggt ggcaccttcg cgcctgtctc gctgctttcg ataagtctct
     3121 agccatttaa aatttttgat gacctgctgc gacgcttttt ttctggcaag atagtcttgt
     3181 aaatgcgggc caagatctgc acactggtat ttcggttttt ggggccgcgg gcggcgacgg
     3241 ggcccgtgcg tcccagcgca catgttcggc gaggcggggc ctgcgagcgc ggccaccgag
     3301 aatcggacgg gggtagtctc aagctggccg gcctgctctg gtgcctggcc tcgcgccgcc
     3361 gtgtatcgcc ccgccctggg cggcaaggct ggcccggtcg gcaccagttg cgtgagcgga
     3421 aagatggccg cttcccggcc ctgctgcagg gagctcaaaa tggaggacgc ggcgctcggg
     3481 agagcgggcg ggtgagtcac ccacacaaag gaaaagggcc tttccgtcct cagccgtcgc
     3541 ttcatgtgac tccacggagt accgggcgcc gtccaggcac ctcgattagt tctcgagctt
     3601 ttggagtacg tcgtctttag gttgggggga ggggttttat gcgatggagt ttccccacac
     3661 tgagtgggtg gagactgaag ttaggccagc ttggcacttg atgtaattct ccttggaatt
     3721 tgcccttttt gagtttggat cttggttcat tctcaagcct cagacagtgg ttcaaagttt
     3781 ttttcttcca tttcaggtgt cgtgagcggc cgctgaactg aattcatcga cgttaactat
     3841 tctagagtac ccatggtgcg caggttcttg gtgaccctcc ggattcggcg cgcgtgcggc
     3901 ccgccgcgag tgagggtttt cgtggttcac atcccgcggc tcacggggga gtgggcagcg
     3961 ccaggggcgc ccgccgctgt ggccctcgtg ctgatgctac tgaggagcca gcgtctaggg
     4021 cagcagccgc ttcctagaag accaggtcat gatgatgggc agcgcccgag tggcggagct
     4081 gctgctgctc cacggcgcgg agcccaactg cgccgacccc gccactctca cccgacccgt
     4141 gcacgacgct gcccgggagg gcttcctgga cacgctggtg gtgctgcacc gggccggggc
     4201 gcggctggac gtgcgcgatg cctggggccg tctgcccgtg gacctggctg agggctagga
     4261 tccgcccctc tccctccccc ccccctaacg ttactggccg aagccgcttg gaataaggcc
     4321 ggtgtgcgtt tgtctatatg ttattttcca ccatattgcc gtcttttggc aatgtgaggg
     4381 cccggaaacc tggccctgtc ttcttgacga gcattcctag gggtctttcc cctctcgcca
     4441 aaggaatgca aggtctgttg aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa
     4501 gacaaacaac gtctgtagcg accctttgca ggcagcggaa ccccccacct ggcgacaggt
     4561 gcctctgcgg ccaaaagcca cgtgtataag atacacctgc aaaggcggca caaccccagt
     4621 gccacgttgt gagttggata gttgtggaaa gagtcaaatg gctctcctca agcgtattca
     4681 acaaggggct gaaggatgcc cagaaggtac cccattgtat gggatctgat ctggggcctc
     4741 ggtacacatg ctttacatgt gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc
     4801 acggggacgt ggttttcctt tgaaaaacac gatgataata tggatgccag agccagcgaa
     4861 gtctgctccc gccccgaaaa agggctccaa gaaggcggtg actaaggcgc agaagaaagg
     4921 cggcaagaag cgcaagcgca gccgcaagga gagctattcc atctatgtgt acaaggttct
     4981 gaagcaggtc caccctgaca ccggcatttc gtccaaggcc atgggcatca tgaattcgtt
     5041 tgtgaacgac attttcgagc gcatcgcagg tgaggcttcc cgcctggcgc attacaacaa
     5101 gcgctcgacc atcacctcca gggagatcca gacggccgtg cgtctgctgc tgcctgggga
     5161 gttggccaag cacgccgtgt ccgagggtac taaggccatc accaagtaca ccagcgctaa
     5221 ggatccaccg gtcgccacca tggtgagcaa gggcgaggag aataacatgg ccatcatcaa
     5281 ggagttcatg cgcttcaagg tgcgcatgga gggctccgtg aacggccacg agttcgagat
     5341 cgagggcgag ggcgagggcc gcccctacga gggctttcag accgctaagc tgaaggtgac
     5401 caagggtggc cccctgccct tcgcctggga catcctgtcc cctcatttca cctacggctc
     5461 caaggcctac gtgaagcacc ccgccgacat ccccgactac ttcaagctgt ccttccccga
     5521 gggcttcaag tgggagcgcg tgatgaacta cggggacggc ggcgtggtga ccgtgaccca
     5581 ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt
     5641 cccctccgac ggccccgtga tgcagaagaa gaccatgggc tgggaggcct cctccgagcg
     5701 gatgtacccc gaggacggtg ccctgaaggg caagatcaag atgaggctga agctgaagga
     5761 cggcggccac tacacctccg aggtcaagac cacctacaag gccaagaagc ccgtgcagct
     5821 gcccggcgcc tacatcgtcg acatcaagtt ggacatcacc tcccacaacg aggactacac
     5881 catcgtggaa cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct
     5941 gtacaagtaa gcggccgcga ctctagacga taccgtcgac ctcgatcgag acctagaaaa
     6001 acatggagca atcacaagta gcaatacagc agctaccaat gctgattgtg cctggctaga
     6061 agcacaagag gaggaggagg tgggttttcc agtcacacct caggtacctt taagaccaat
     6121 gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg
     6181 gctaattcac tcccaacgaa gacaagatat ccttgatctg tggatctacc acacacaagg
     6241 ctacttccct gattggcaga actacacacc agggccaggg atcagatatc cactgacctt
     6301 tggatggtgc tacaagctag taccagttga gcaagagaag gtagaagaag ccaatgaagg
     6361 agagaacacc cgcttgttac accctgtgag cctgcatggg atggatgacc cggagagaga
     6421 agtattagag tggaggtttg acagccgcct agcatttcat cacatggccc gagagctgca
     6481 tccggactgt actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta
     6541 actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg
     6601 tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg
     6661 gaaaatctct agcagcatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     6721 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     6781 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     6841 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     6901 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     6961 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     7021 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     7081 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     7141 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     7201 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     7261 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     7321 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     7381 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     7441 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     7501 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     7561 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     7621 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     7681 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     7741 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     7801 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     7861 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     7921 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     7981 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     8041 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     8101 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     8161 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     8221 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     8281 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     8341 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     8401 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     8461 cacatttccc cgaaaagtgc cacct
//
