LOCUS       sequence-436345-        6180 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  C terminal fusion of EGFP to mouse HP1-gamma (Cbx3) containing M158V
            mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6180)
  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 6180)
  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-gamma-M158V-EGFP
FEATURES             Location/Qualifiers
     source          1..6180
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     1678..1700
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1762..1781
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2389..2398
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2395..2397,2398..2400,2401..3114)
                     /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(2395..2397,2398..2400,2401..3111)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2440..2461)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2701..2720)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3048..3069
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3201..3220)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3266..3321
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3320..3339)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3682..3698)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3682..3698)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3695..3717)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3706..3722
                     /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(3730..3736,3737..3754,3755..3760))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3775..3796
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3855..4050
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3901..4036
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4056..4190
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4093..4112)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4147..4166
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4258..4275)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4429..5017)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4509..4528)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5188..5979,5980..6048))
                     /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     5811..5830
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6049..6153)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca
      421 tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag
      481 cgatgggggc gggggggggg ggggggggcg ggcgcggggg gggggggggg gggggggggg
      541 gggggggggg gggggggggg gggggggggg gggggggcgg ggcggggcgg agaggtgcgg
      601 cggcagccaa tcagagcggc gcgctccgaa agtttccttt tatggcgagg cggcggcggc
      661 ggcggcccta taaaaagcga agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc
      721 ccgtgccccg ctccgccgcc gcctcgcgcc gcccgccccg gctctgactg accgcgttac
      781 tcccacaggt gagcgggcgg gacggccctt ctcctccggg ctgtaattag cgcttggttt
      841 aatgacggct tgtttctttt ctgtggctgc gtgaaagcct tgaggggctc cgggagggcc
      901 ctttgtgcgg ggggagcggc tcggggggtg cgtgcgtgtg tgtgtgcgtg gggagcgccg
      961 cgtgcggctc cgcgctgccc ggcggctgtg agcgctgcgg gcgcggcgcg gggctttgtg
     1021 cgctccgcag tgtgcgcgag gggagcgcgg ccgggggcgg tgccccgcgg tgcggggggg
     1081 gctgcgaggg gaacaaaggc tgcgtgcggg gtgtgtgcgt gggggggtga gcagggggtg
     1141 tgggcgcgtc ggtcgggctg caaccccccc tgcacccccc tccccgagtt gctgagcacg
     1201 gcccggcttc gggtgcgggg ctccgtacgg ggcgtggcgc ggggctcgcc gtgccgggcg
     1261 gggggtggcg gcaggtgggg gtgccgggcg gggcggggcc gcctcgggcc gggggggggt
     1321 cggggggggg gggcgggggg ccccgggggg cggggggggg cgggggggag gggcgcggcg
     1381 gcccccggag cgccggcggc tgtcgaggcg cggcgagccg cagccattgc cttttatggt
     1441 aatcgtgcga gagggcgcag ggacttcctt tgtcccaaat ctgtgcggag ccgaaatctg
     1501 ggaggcgccg ccgcaccccc tctagcgggc gcggggcgaa gcggtgcggc gccggcagga
     1561 aggaaatggg cggggagggc cttcgtgcgt cgccgcgccg ccgtcccctt ctccctctcc
     1621 agcctcgggg ctgtccgcgg ggggacggct gccttcgggg gggacggggc agggcggggt
     1681 tcggcttctg gcgtgtgacc ggcggctcta gagcctctgc taaccatgtt catgccttct
     1741 tctttttcct acagctcctg ggcaacgtgc tggttattgt gctgtctcat cattttggca
     1801 aagaattctg cagtcgacgg tacctaaaaa atggcctcca ataaaactac attgcaaaaa
     1861 atgggaaaga aacaaaatgg aaagagtaaa aaagttgaag aggcagagcc tgaagaattt
     1921 gtggtagaaa aagtactgga ccgtcgtgta gtgaatggga aggtggagta tttcctgaag
     1981 tggaaggggt tcacagatgc tgataatact tgggaaccag aagaaaattt agattgtcca
     2041 gaattaattg aagcatttct taattctcaa aaagctggta aagaaaaaga tggtacaaaa
     2101 aggaaatctt tatctgacag tgaatctgat gatagcaaat cgaagaagaa gagagatgct
     2161 gctgacaaac caaggggctt tgccagaggt ctcgaccctg aacgaataat cggcgccaca
     2221 gacagcagcg gagagttaat gtttctcatg aagtggaagg actcggacga ggccgacttg
     2281 gtgctggcaa aggaggcgaa cgtgaagtgt cctcagattg tcattgcctt ctacgaggag
     2341 cggctgactt ggcattcttg tcctgaagat gaagcacaat taccggtcgc caccatggtg
     2401 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     2461 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     2521 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     2581 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     2641 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     2701 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     2761 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     2821 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc
     2881 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     2941 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     3001 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     3061 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaaagcggc
     3121 cgcactcctc aggtgcaggc tgcctatcag aaggtggtgg ctggtgtggc caatgccctg
     3181 gctcacaaat accactgaga tctttttccc tctgccaaaa attatgggga catcatgaag
     3241 ccccttgagc atctgacttc tggctaataa aggaaattta ttttcattgc aatagtgtgt
     3301 tggaattttt tgtgtctctc actcggaagg acatatggga gggcaaatca tttaaaacat
     3361 cagaatgagt atttggttta gagtttggca acatatgccc atatgctggc tgccatgaac
     3421 aaaggttggc tataaagagg tcatcagtat atgaaacagc cccctgctgt ccattcctta
     3481 ttccatagaa aagccttgac ttgaggttag atttttttta tattttgttt tgtgttattt
     3541 ttttctttaa catccctaaa attttcctta catgttttac tagccagatt tttcctcctc
     3601 tcctgactac tcccagtcat agctgtccct cttctcttat ggagatccct cgacctgcag
     3661 cccaagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     3721 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     3781 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     3841 gtgccagcgg atccgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     3901 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     3961 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     4021 ggcttttttg gaggcctagg cttttgcaaa aagctaactt gtttattgca gcttataatg
     4081 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     4141 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc cgctgcatta
     4201 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     4261 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     4321 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     4381 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     4441 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     4501 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     4561 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     4621 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     4681 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     4741 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     4801 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     4861 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     4921 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     4981 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     5041 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     5101 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     5161 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     5221 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     5281 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     5341 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     5401 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     5461 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     5521 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     5581 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     5641 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     5701 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     5761 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     5821 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     5881 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     5941 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     6001 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     6061 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     6121 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctgg
//
