LOCUS       sequence-436311-        6123 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  C terminal fusion of EGFP to mouse HP1-alpha (Cbx5) containing F44E 
            mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6123)
  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 6123)
  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-alpha-F44E-EGFP
FEATURES             Location/Qualifiers
     source          1..6123
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          657..1673
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1597..1619
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1681..1700
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2332..2341
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2338..2340,2341..2343,2344..3057)
                     /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(2338..2340,2341..2343,2344..3054)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2383..2404)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2644..2663)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2991..3012
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3144..3163)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3209..3264
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3263..3282)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3625..3641)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3625..3641)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3638..3660)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3649..3665
                     /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(3673..3679,3680..3697,3698..3703))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3718..3739
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3798..3993
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3844..3979
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3999..4133
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4036..4055)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4090..4109
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4201..4218)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4372..4960)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4452..4471)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5131..5922,5923..5991))
                     /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     5754..5773
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5992..6096)
                     /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 gggggggggg gggggggggg cgcgcggggg ggggggcggg gcgggggggg gggggggggg
      541 gggggaggcg gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt
      601 ttatggcgag gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag
      661 tcgctgcgcg ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc
      721 ggctctgact gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctcctccgg
      781 gctgtaatta gcgcttggtt taatgacggc ttgtttcttt tctgtggctg cgtgaaagcc
      841 ttgaggggct ccgggagggc cctttgtgcg gggggagcgg ctcggggggt gcgtgcgtgt
      901 gtgtgtgcgt ggggagcgcc gcgtgcggct ccgcgctgcc cggcggctgt gagcgctgcg
      961 ggcgcggcgc ggggctttgt gcgctccgca gtgtgcgcga ggggagcgcg gccgggggcg
     1021 gtgccccgcg gtgcgggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgt cggtcgggct gcaacccccc ctgcaccccc
     1141 ctccccgagt tgctgagcac ggcccggctt cgggtgcggg gctccgtacg gggcgtggcg
     1201 cggggctcgc cgtgccgggc ggggggtggc ggcaggtggg ggtgccgggc ggggcggggc
     1261 cgcctcgggc cggggagggc tcgggggagg ggcgcggcgg cccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tgtgcggagc cgaaatctgg gaggcgccgc cgcaccccct
     1441 ctagcgggcg cggggcgaag cggtgcggcg ccggcaggaa ggaaatgggc ggggagggcc
     1501 ttcgtgcgtc gccgcgccgc cgtccccttc tccctctcca gcctcggggc tgtccgcggg
     1561 gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg cgtgtgaccg
     1621 gcggctctag agcctctgct aaccatgttc atgccttctt ctttttccta cagctcctgg
     1681 gcaacgtgct ggttattgtg ctgtctcatc attttggcaa agaattctgc agtcgacggt
     1741 acccacgaca tgggaaagaa gaccaagagg acagccgaca gctcttcttc agaggatgag
     1801 gaggaatatg tggtggaaaa ggtgttggac aggcgcatgg ttaaggggca agtggaatat
     1861 ctgttgaagt ggaaaggcga gtctgaggag cacaatactt gggaacctga gaagaacttg
     1921 gattgtcctg aactaatttc tgagtttatg aaaaagtata agaagatgaa ggagggtgaa
     1981 aacaataagc ccagggagaa atcagaagga aacaagagga aatccagttt ctccaacagc
     2041 gctgatgata ttaaatctaa aaaaaagaga gagcaaagca atgatatcgc tcggggcttt
     2101 gagagaggac tggaaccaga aaagatcatc ggagcaacag attcctgcgg tgacttaatg
     2161 ttcttaatga aatggaaaga cacagatgaa gctgacctgg ttcttgcaaa agaagctaac
     2221 gtgaagtgtc cacagattgt gatagcattt tatgaagaga gactgacgtg gcacgcatat
     2281 ccagaggatg cggaaaacaa agaaaaagaa agcgcgaaga gcttaccggt cgccaccatg
     2341 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     2401 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     2461 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     2521 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     2581 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     2641 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     2701 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     2761 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     2821 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     2881 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     2941 ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     3001 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaaagc
     3061 ggccgcactc ctcaggtgca ggctgcctat cagaaggtgg tggctggtgt ggccaatgcc
     3121 ctggctcaca aataccactg agatcttttt ccctctgcca aaaattatgg ggacatcatg
     3181 aagccccttg agcatctgac ttctggctaa taaaggaaat ttattttcat tgcaatagtg
     3241 tgttggaatt ttttgtgtct ctcactcgga aggacatatg ggagggcaaa tcatttaaaa
     3301 catcagaatg agtatttggt ttagagtttg gcaacatatg cccatatgct ggctgccatg
     3361 aacaaaggtt ggctataaag aggtcatcag tatatgaaac agccccctgc tgtccattcc
     3421 ttattccata gaaaagcctt gacttgaggt tagatttttt ttatattttg ttttgtgtta
     3481 tttttttctt taacatccct aaaattttcc ttacatgttt tactagccag atttttcctc
     3541 ctctcctgac tactcccagt catagctgtc cctcttctct tatggagatc cctcgacctg
     3601 cagcccaagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     3661 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     3721 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     3781 gtcgtgccag cggatccgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     3841 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3901 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     3961 ggaggctttt ttggaggcct aggcttttgc aaaaagctaa cttgtttatt gcagcttata
     4021 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     4081 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg atccgctgca
     4141 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     4201 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     4261 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     4321 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     4381 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     4441 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     4501 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     4561 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     4621 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     4681 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     4741 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     4801 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     4861 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     4921 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     4981 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     5041 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     5101 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     5161 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     5221 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     5281 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     5341 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     5401 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     5461 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     5521 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     5581 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     5641 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     5701 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     5761 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     5821 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     5881 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     5941 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     6001 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     6061 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     6121 tgg
//
