LOCUS       sequence-436321-        6109 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  C terminal fusion of EGFP to mouse HP1-beta (Cbx1) containing I161E 
            mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6109)
  AUTHORS   Newman AG, Sharif J, Bessa P, Zaqout S, Brown JP, Richter D, 
            Dannenberg R, Nakayama M, Mueller S, Schaub T, Manickaraj S, 
            Boehm-Sturm P, Ohara O, Koseki H, Singh PB, Tarabykin V
  TITLE     Glial reactivity and cognitive decline follow chronic 
            heterochromatin loss in neurons.
  JOURNAL   Nat Commun. 2025 Aug 8;16(1):7325. doi: 10.1038/s41467-025-61319-7.
  PUBMED    40781223
REFERENCE   2  (bases 1 to 6109)
  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: "2025-08-8"; volume: "16(1):7325. doi"; pages: " 
            10.1038/s41467-025-61319-7"
COMMENT     Sequence Label: pCAG-HP1-beta-I161E-EGFP
FEATURES             Location/Qualifiers
     source          1..6109
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          662..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1602..1624
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1686..1705
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2319..2328
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2325..2327,2328..2330,2331..3044)
                     /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(2325..2327,2328..2330,2331..3041)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2370..2391)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2631..2650)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2978..2999
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3131..3150)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3196..3251
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3250..3269)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3612..3628)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3612..3628)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3625..3647)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3636..3652
                     /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(3660..3666,3667..3684,3685..3690))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3705..3726
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3785..3980
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3831..3966
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3986..4120
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4023..4042)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4077..4096
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4188..4205)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4359..4947)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4439..4458)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5118..5909,5910..5978))
                     /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     5741..5760
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5979..6083)
                     /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 gggggggcgc gccrggsggg gsgggggggg gsgrggggsg
      541 gggsggggsg rggcggagag gkgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
      721 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
      781 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
      841 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg
      901 cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg
      961 ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt
     1081 gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gkgggggtgc cgggcggggc
     1261 ggggccgcct cgggccgggg rgggctcggg ggrggggcgc ggcggccccc ggagcgccgg
     1321 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     1381 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     1441 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     1501 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     1561 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1621 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tctgcagtcg
     1741 acggtacccg tactatgggg aaaaagcaaa acaagaagaa agtggaggag gtactagaag
     1801 aagaggaaga ggaatatgtg gtggaaaaag ttcttgatcg gcgagttgtc aagggcaagg
     1861 tggaatatct tctaaagtgg aagggtttct cagatgagga caacacttgg gagccagaag
     1921 agaatctgga ttgccctgac cttattgctg agtttctaca gtcacagaaa acagctcatg
     1981 agacagataa gtcagaggga ggcaagcgca aagctgattc tgattctgaa gataaaggag
     2041 aggaaagcaa accaaagaag aagaaagaag agtcagaaaa gccacgaggc tttgcccggg
     2101 gtttggagcc agagcggatt attggagcta ctgactccag tggagagctc atgttcctga
     2161 tgaaatggaa aaactctgat gaggctgacc tggtccctgc caaggaagcc aatgtcaagt
     2221 gcccacaggt tgtcgaatcc ttctatgagg aaaggctaac gtggcattcc tacccctcag
     2281 aggatgatga caaaaaagac gacaagaatt taccggtcgc caccatggtg agcaagggcg
     2341 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     2401 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     2461 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga
     2521 cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca
     2581 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     2641 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     2701 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     2761 acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact
     2821 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     2881 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt
     2941 ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     3001 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaaagcggc cgcactcctc
     3061 aggtgcaggc tgcctatcag aaggtggtgg ctggtgtggc caatgccctg gctcacaaat
     3121 accactgaga tctttttccc tctgccaaaa attatgggga catcatgaag ccccttgagc
     3181 atctgacttc tggctaataa aggaaattta ttttcattgc aatagtgtgt tggaattttt
     3241 tgtgtctctc actcggaagg acatatggga gggcaaatca tttaaaacat cagaatgagt
     3301 atttggttta gagtttggca acatatgccc atatgctggc tgccatgaac aaaggttggc
     3361 tataaagagg tcatcagtat atgaaacagc cccctgctgt ccattcctta ttccatagaa
     3421 aagccttgac ttgaggttag atttttttta tattttgttt tgtgttattt ttttctttaa
     3481 catccctaaa attttcctta catgttttac tagccagatt tttcctcctc tcctgactac
     3541 tcccagtcat agctgtccct cttctcttat ggagatccct cgacctgcag cccaagcttg
     3601 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     3661 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     3721 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagcgg
     3781 atccgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     3841 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     3901 cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg
     3961 gaggcctagg cttttgcaaa aagctaactt gtttattgca gcttataatg gttacaaata
     4021 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     4081 tttgtccaaa ctcatcaatg tatcttatca tgtctggatc cgctgcatta atgaatcggc
     4141 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     4201 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     4261 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     4321 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     4381 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     4441 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     4501 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     4561 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4621 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     4681 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     4741 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     4801 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     4861 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     4921 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     4981 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     5041 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     5101 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     5161 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     5221 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     5281 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     5341 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     5401 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     5461 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     5521 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     5581 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     5641 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     5701 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     5761 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     5821 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     5881 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     5941 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     6001 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     6061 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctg
//
