LOCUS       sequence-388458-        5191 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To over express mCherry-CAAX under gfaABC1D promoter in astrocytes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5191)
  AUTHORS   Bindu DS, Tan CX, Savage JT, Eroglu C
  TITLE     GEARBOCS: An Adeno Associated Virus Tool for In Vivo Gene Editing in
            Astrocytes.
  JOURNAL   bioRxiv. 2023 Jan 19:2023.01.17.524433. doi: 
            10.1101/2023.01.17.524433. Preprint.
  PUBMED    36711516
REFERENCE   2  (bases 1 to 5191)
  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: "bioRxiv. 
            2023 Jan 19:2023.01.17.524433. doi: 10.1101/2023.01.17.524433. 
            Preprint."
COMMENT     Sequence Label: pgfaABC1D-mCherry-CAAX
FEATURES             Location/Qualifiers
     source          1..5191
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      908..1496
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1397..1416
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    1784..1805
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1820..1825,1826..1843,1844..1850)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1858..1874
                     /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)."
     primer_bind     1863..1885
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1882..1898
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1882..1898
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   1936..2065
                     /label=AAV2 ITR
     intron          2905..3037
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     3020..3039
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        3082..3100
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      3138..3147
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3144..3851
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(3289..3307)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3557..3576
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    3893..4014
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3904..3923)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3958..3977
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4176..4305
                     /label=AAV2 ITR
     primer_bind     complement(4324..4341)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4324..4340)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4333..4355)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4482..4937
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4569..4588)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4779..4800
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4963..5067
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5068..5136,5137..5191,1..737)
                     /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"
ORIGIN
        1 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
       61 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
      121 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
      181 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
      241 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
      301 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
      361 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
      421 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
      481 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
      541 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
      601 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
      661 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
      721 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
      781 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
      841 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
      901 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
      961 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     1021 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     1081 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     1141 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     1201 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     1261 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     1321 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     1381 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     1441 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     1501 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     1561 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     1621 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     1681 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     1741 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca
     1801 ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg
     1861 tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc agatttaatt
     1921 aaggccttaa ttaggctgcg cgctcgctcg ctcactgagg ccgcccgggc aaagcccggg
     1981 cgtcgggcga cctttggtcg cccggcctca gtgagcgagc gagcgcgcag agagggagtg
     2041 gccaactcca tcactagggg ttccttgtag ttaatgatta acccgccatg ctacttatct
     2101 acgtagccat gctctaggaa gatctaacat atcctggtgt ggagtagggg acgctgctct
     2161 gacagaggct cgggggcctg agctggctct gtgagctggg gaggaggcag acagccaggc
     2221 cttgtctgca agcagacctg gcagcattgg gctggccgcc ccccagggcc tcctcttcat
     2281 gcccagtgaa tgactcacct tggcacagac acaatgttcg gggtgggcac agtgcctgct
     2341 tcccgccgca ccccagcccc cctcaaatgc cttccgagaa gcccattgag cagggggctt
     2401 gcattgcacc ccagcctgac agcctggcat cttgggataa aagcagcaca gccccctagg
     2461 ggctgccctt gctgtgtggc gccaccggcg gtggagaaca aggctctatt cagcctgtgc
     2521 ccaggaaagg ggatcagggg atgcccaggc atggacagtg ggtggcaggg ggggagagga
     2581 gggctgtctg cttcccagaa gtccaaggac acaaatgggt gaggggagag ctctccccat
     2641 agctgggctg cggcccaacc ccaccccctc aggctatgcc agggggtgtt gccaggggca
     2701 cccgggcatc gccagtctag cccactcctt cataaagccc tcgcatccca ggagcgagca
     2761 gagccagagc aggttggaga ggagacgcat cacctccgct gctcgcaagc tttattgcgg
     2821 tagtttatca cagttaaatt gctaacgcct gacacaacag tctcgaactt aagctgcaga
     2881 agttggtcgt gaggcactgg gcaggtaagt atcaaggtta caagacaggt ttaaggagac
     2941 caatagaaac tgggcttgtc gagacagaga agactcttgc gtttctgata ggcacctatt
     3001 ggtcttactg acatccactt tgcctttctc tccacaggtg tccactccca gttcaattac
     3061 agctcttaag gctagagtac ttaatacgac tcactatagg ctagcctcga gctcaagctt
     3121 cgaattctgc agtcgacgcc accatggtga gcaagggcga ggaggataac atggccatca
     3181 tcaaggagtt catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg
     3241 agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg
     3301 tgaccaaggg tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg
     3361 gctccaaggc ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc
     3421 ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga
     3481 cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca
     3541 acttcccctc cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctcctccg
     3601 agcggatgta ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga
     3661 aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc
     3721 agctgcccgg cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact
     3781 acaccatcgt ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg
     3841 agctgtacaa gtgctgcatc atcctgtaag cggccgcttc gagcagacat gataagatac
     3901 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     3961 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
     4021 aacaattgca ttcattttat gtttcaggtt cagggggaga tgtgggaggt tttttaaagc
     4081 aagtaaaacc tctacaaatg tggtaaaatc gataaggatc ttcctagagc atggctacgt
     4141 agataagtag catggcgggt taatcattaa ctacaaggaa cccctagtga tggagttggc
     4201 cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg
     4261 cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagcctta attaacctaa
     4321 ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
     4381 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
     4441 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc
     4501 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     4561 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     4621 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     4681 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     4741 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     4801 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     4861 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat
     4921 attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     4981 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     5041 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     5101 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     5161 aaagatgctg aagatcagtt gggtgcacga g
//
