LOCUS       sequence-457035-        6592 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression CDKN1A in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6592)
  AUTHORS   Lin L, Zhao J, Kubota N, Li Z, Lam YL, Nguyen LP, Yang L, Pokharel 
            SP, Blue SM, Yee BA, Chen R, Yeo GW, Chen CW, Chen L, Zheng S
  TITLE     Epistatic interactions between NMD and TRP53 control progenitor cell
            maintenance and brain size.
  JOURNAL   Neuron. 2024 Jul 3;112(13):2157-2176.e12. doi: 
            10.1016/j.neuron.2024.04.006. Epub 2024 May 1.
  PUBMED    38697111
REFERENCE   2  (bases 1 to 6592)
  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.neuron.2024.04.006"; journalName: "Neuron"; date: 
            "2024-07-3- 3"; volume: "112"; issue: "13"; pages: "2157-2176.e12"
COMMENT     Sequence Label: pCAGIG-CDKN1A
FEATURES             Location/Qualifiers
     source          1..6592
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..662
                     /label=chicken beta-actin promoter
     intron          663..1679
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1603..1625
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1687..1706
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     misc_feature    join(2244..2818,2819..2821,2822..2830)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2411..2428)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2638..2657
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2831..2833,2834..2836,2837..3550)
                     /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             join(2831..2833,2834..2836,2837..3547)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2876..2897)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3137..3156)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3484..3505
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3614..3633)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3679..3734
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3733..3752)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4095..4111)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4095..4111)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4108..4130)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4119..4135
                     /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(4143..4149,4150..4167,4168..4173))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4188..4209
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4268..4463
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4314..4449
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4469..4603
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4506..4525)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4560..4579
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4671..4688)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4842..5430)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4922..4941)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5601..6392,6393..6461))
                     /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     6224..6243
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6462..6566)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg ggggggcgcg ggccaggcgg ggcggggcgg ggcgaggggc
      541 ggggcggggc gaggcggaga ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt
      601 ttccttttat ggcgaggcgg cggcggcggc ggccctataa aaagcgaagc gcgcggcggg
      661 cgggagtcgc tgcgcgctgc cttcgccccg tgccccgctc cgccgccgcc tcgcgccgcc
      721 cgccccggct ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc
      781 ctccgggctg taattagcgc ttggtttaat gacggcttgt ttcttttctg tggctgcgtg
      841 aaagccttga ggggctccgg gagggccctt tgtgcggggg gagcggctcg gggggtgcgt
      901 gcgtgtgtgt gtgcgtgggg agcgccgcgt gcggctccgc gctgcccggc ggctgtgagc
      961 gctgcgggcg cggcgcgggg ctttgtgcgc tccgcagtgt gcgcgagggg agcgcggccg
     1021 ggggcggtgc cccgcggtgc ggggggggct gcgaggggaa caaaggctgc gtgcggggtg
     1081 tgtgcgtggg ggggtgagca gggggtgtgg gcgcgtcggt cgggctgcaa ccccccctgc
     1141 acccccctcc ccgagttgct gagcacggcc cggcttcggg tgcggggctc cgtacggggc
     1201 gtggcgcggg gctcgccgtg ccgggcgggg ggtggcggca ggtgggggtg ccgggcgggg
     1261 cggggccgcc tcgggccggg gagggctcgg gggaggggcg cggcggcccc cggagcgccg
     1321 gcggctgtcg aggcgcggcg agccgcagcc attgcctttt atggtaatcg tgcgagaggg
     1381 cgcagggact tcctttgtcc caaatctgtg cggagccgaa atctgggagg cgccgccgca
     1441 ccccctctag cgggcgcggg gcgaagcggt gcggcgccgg caggaaggaa atgggcgggg
     1501 agggccttcg tgcgtcgccg cgccgccgtc cccttctccc tctccagcct cggggctgtc
     1561 cgcgggggga cggctgcctt cgggggggac ggggcagggc ggggttcggc ttctggcgtg
     1621 tgaccggcgg ctctagagcc tctgctaacc atgttcatgc cttcttcttt ttcctacagc
     1681 tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt tggcaaagaa ttcatgtcca
     1741 atcctggtga tgtccgacct gttccgcaca ggagcaaagt gtgccgttgt ctcttcggtc
     1801 ccgtggacag tgagcagttg cgccgtgatt gcgatgcgct catggcgggc tgtctccagg
     1861 aggcccgaga acggtggaac tttgacttcg tcacggagac gccgctggag ggcaactttg
     1921 tctgggagcg cgttcggagc ctagggctgc ccaaggtcta cctgagccct gggtcccgca
     1981 gccgtgacga cctgggaggg gacaagaggc ccagtacttc ctctgccctg ctgcaggggc
     2041 cagctccaga ggaccacgtg gccttgtcgc tgtcttgcac tctggtgtct gagcggcctg
     2101 aagattcccc gggtgggccc ggaacatctc agggccgaaa acggaggcag accagcctga
     2161 cagatttcta tcactccaag cgcagattgg tcttctgcaa gagaaaaccc tgagcggccg
     2221 ctacgtaaat tccgcccccc ccccccctct ccctcccccc cccctaacgt tactggccga
     2281 agccgcttgg aataaggccg gtgtgcgttt gtctatatgt tattttccac catattgccg
     2341 tcttttggca atgtgagggc ccggaaacct ggccctgtct tcttgacgag cattcctagg
     2401 ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa ggaagcagtt
     2461 cctctggaag cttcttgaag acaaacaacg tctgtagcga ccctttgcag gcagcggaac
     2521 cccccacctg gcgacaggtg cctctgcggc caaaagccac gtgtataaga tacacctgca
     2581 aaggcggcac aaccccagtg ccacgttgtg agttggatag ttgtggaaag agtcaaatgg
     2641 ctctcctcaa gcgtattcaa caaggggctg aaggatgccc agaaggtacc ccattgtatg
     2701 ggatctgatc tggggcctcg gtgcacatgc tttacatgtg tttagtcgag gttaaaaaaa
     2761 cgtctaggcc ccccgaacca cggggacgtg gttttccttt gaaaaacacg atgataatat
     2821 ggccacaacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     2881 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     2941 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     3001 ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga
     3061 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     3121 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     3181 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     3241 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa
     3301 gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt
     3361 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     3421 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     3481 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     3541 gtacaagtaa agcggccggc cgccagcaca gtggtcgatc gaccaatgcc ctggctcaca
     3601 aataccactg agatcttttt ccctctgcca aaaattatgg ggacatcatg aagccccttg
     3661 agcatctgac ttctggctaa taaaggaaat ttattttcat tgcaatagtg tgttggaatt
     3721 ttttgtgtct ctcactcgga aggacatatg ggagggcaaa tcatttaaaa catcagaatg
     3781 agtatttggt ttagagtttg gcaacatatg cccatatgct ggctgccatg aacaaaggtt
     3841 ggctataaag aggtcatcag tatatgaaac agccccctgc tgtccattcc ttattccata
     3901 gaaaagcctt gacttgaggt tagatttttt ttatattttg ttttgtgtta tttttttctt
     3961 taacatccct aaaattttcc ttacatgttt tactagccag atttttcctc ctctcctgac
     4021 tactcccagt catagctgtc cctcttctct tatggagatc cctcgacctg cagcccaagc
     4081 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     4141 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     4201 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     4261 cggatccgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     4321 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     4381 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     4441 ttggaggcct aggcttttgc aaaaagctaa cttgtttatt gcagcttata atggttacaa
     4501 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     4561 tggtttgtcc aaactcatca atgtatctta tcatgtctgg atccgctgca ttaatgaatc
     4621 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     4681 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     4741 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     4801 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     4861 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     4921 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     4981 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5041 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     5101 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5161 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     5221 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     5281 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     5341 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     5401 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     5461 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     5521 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     5581 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     5641 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     5701 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     5761 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     5821 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     5881 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     5941 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6001 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6061 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     6121 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     6181 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     6241 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     6301 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     6361 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     6421 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     6481 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     6541 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tg
//
