LOCUS       sequence-465035-        5515 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV production plasmid for PGK(synEPOR) vector from Figs. 3-5 that 
            mediates HDR at CCR5 locus using CCR5 gRNA. synEPOR is followed by 
            BGH polyA. AAV genome is flanked by AAV2 ITRs..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5515)
  AUTHORS   Shah AP, Majeti KR, Ekman FK, Selvaraj S, Soupene E, Chati P, Sinha 
            R, Luna SE, Charlesworth CT, McCreary T, Lesch BJ, Tran T, Sharma D,
            Chu SN, Porteus MH, Cromer MK
  TITLE     Engineering inducible signaling receptors to enable 
            erythropoietin-free erythropoiesis
  JOURNAL   bioRxiv 2024.04.12.589260
REFERENCE   2  (bases 1 to 5515)
  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 
            2024.04.12.589260"
COMMENT     Sequence Label: MKC177_CCR5(PGK-synEPOR)
FEATURES             Location/Qualifiers
     source          1..5515
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(602..922)
                     /codon_start=1
                     /product="F36V mutant of FK506-binding protein FKBP12"
                     /label=FKBP-V
                     /note="binds synthetic ligands that do not bind wild-type 
                     FKBP"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     promoter        complement(1083..1593)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1146..1164)
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     protein_bind    1716..1727
                     /label=ZF binding site
                     /bound_moiety="CCR5TC zinc-finger domain"
                     /note="target sequence for the CCR5TC zinc-finger domain 
                     from the CCR5-224 (+) zinc-finger nuclease (Perez et al., 
                     2008; Gross et al., 2013)"
     repeat_region   2031..2171
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2031..2160
                     /label=AAV2 ITR
     rep_origin      2246..2701
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2333..2352)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2543..2564
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2718..2737)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2837..2859
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2897..2915)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2983..3087
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3088..3156,3157..3948)
                     /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     complement(3306..3325)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4119..4707
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4608..4627
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   4769..4898
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     polyA_signal    join(5307..5515,1..16)
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 caactagaag gcacagttat ggacgcaaga gctgggagct ggggtccagg atagtgtact
       61 caaagctggc agcagaggct ccctctgggc tgggcttcga ggccaaagca gatgagcagg
      121 aggatgcttc tgagccttca tccatggcca ctatgtccac actgccacca ggccctggga
      181 ggtcctcact gggcgggttc cggggcagca accatttgtc cagcaccaga taggtatcct
      241 gggcatgctc actgcccact ggctccagca gggggccctc atcatctgtc cccggctcca
      301 ctgcctgcat cgtcccccag cagcgctctg agaggacttc cagggaagca ggtgggtcct
      361 ccgtgaaggg ggtgcagggg ctccaccaca ggcagccatc attctggtac agccacagct
      421 ggaagttacc cttgtgggtg gtgaagaggc cttcaaactc gctctctggg ctcgggatgc
      481 caggccagat cttctgcttc agagcccggc ggtgggagag cagcgcgagc acggtcagca
      541 gcaccaggat gaccacgagg atgagggaga gcgtcaggat gaggggacta cctcctcctc
      601 cttccagttt tagaagctcc acatcgaaga cgagagtggc atgtggtggg atgatgcctg
      661 ggtgcccagt ggcaccatag gcataatctg gagatatagt cagtttggct ctctgaccca
      721 cactcatctg ggcaacccct tcttcccagc ctcggatcac ctcctgcttg cctagcataa
      781 acttaaaggg cttgtttctg tcccgggagg aatcaacttt ctttccatct tcaagcatcc
      841 cggtgtagtg caccacgcag gtctggccgc gcttggggaa ggtgcgcccg tctcctgggg
      901 agatggtttc cacctgcact ccctcaagca tgcttccgcc gccgcctggg gcagggaagg
      961 cagcaggcaa caccaggagc agccccaggg agaaggcaac tggaccgaag gcgcttgtgg
     1021 agaaggagtt catggtggcg atggatcctt aagcttaatt aacctaggtt ttggaaatac
     1081 agctggggag agaggtcggt gattcggtca acgagggagc cgactgccga cgtgcgctcc
     1141 ggaggcttgc agaatgcgga acaccgcgcg ggcaggaaca gggcccacac taccgcccca
     1201 caccccgcct cccgcaccgc cccttcccgg ccgctgctct cggcgcgccc tgctgagcag
     1261 ccgctattgg ccacagccca tcgcggtcgg cgcgctgcca ttgctccctg gcgctgtccg
     1321 tctgcgaggg tactagtgag acgtgcggct tccgtttgtc acgtccggca cgccgcgaac
     1381 cgcaaggaac cttcccgact taggggcgga gcaggaagcg tcgccggggg gcccacaagg
     1441 gtagcggcga agatccgggt gacgctgcga acggacgtga agaatgtgcg agacccaggg
     1501 tcggcgccgc tgcgtttccc ggaaccacgc ccagagcagc cgcgtccctg cgcaaaccca
     1561 gggctgcctt ggaaaaggcg caaccccaac cccgtggaat tccctaggga taacagggta
     1621 atgaagggga cagtaagaag gaaaaacagg tcagagatgg ccaggttgag caggtagatg
     1681 tcagtcatgc tcttcagcct tttgcagttt atcaggatga ggatgaccag catgttgccc
     1741 acaaaaccaa agatgaacac cagtgagtag agcggaggca ggaggcgggc tgcgatttgc
     1801 ttcacattga ttttttggca gggctccgat gtataataat tgatgtcata gattggactt
     1861 gacacttgat aatccatctt gttccaccct gtgcataaat aaaaagtgat cttttataaa
     1921 gtcctagaat gtatttagtt gccctccatg aatgcaaact gttttataca tcaataggtt
     1981 tttaattgcc tacatagatg tctacattga attaactctc ttgcggccgc aggaacccct
     2041 agtgatggag ttggccactc cttctctgcg cgctcgctcg ctcactgagg ccgggcgacc
     2101 aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag
     2161 ctgcctgcag gggcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca
     2221 ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt
     2281 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccttagcgcc cgctcctttc
     2341 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     2401 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     2461 ttgggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     2521 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaactct
     2581 atctcgggct attcttttga tttataaggg attttgccga tttcggtcta ttggttaaaa
     2641 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt
     2701 ttatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
     2761 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     2821 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     2881 cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
     2941 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     3001 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     3061 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     3121 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     3181 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     3241 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     3301 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     3361 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     3421 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     3481 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     3541 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     3601 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     3661 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     3721 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     3781 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     3841 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     3901 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     3961 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     4021 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     4081 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     4141 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     4201 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     4261 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     4321 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     4381 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     4441 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     4501 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     4561 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     4621 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     4681 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     4741 gccttttgct ggccttttgc tcacatgtcc tgcaggcagc tgcgcgctcg ctcgctcact
     4801 gaggccgccc gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg
     4861 cagagaggga gtggccaact ccatcactag gggttcctgc ggccgcttag gattcccgag
     4921 tagcagatga ccatgacaag cagcggcagg accagcccca agatgactat ctttaatgtc
     4981 tggaaattct tccagaattg atactgactg tatggaaaat gagagctgca ggtgtaatga
     5041 agaccttctt tttgagatct ggtaaagatg attcctggga gagacgcaaa cacagccacc
     5101 acccaagtga tcacacttgt caccacccca aaggtgaccg tcctggcttt taaagcaaac
     5161 acagcatgga cgacagccag gtacctatcg attgtcagga ggatgatgaa gaagattcca
     5221 gagaagaagc ctataaaata gagccctgtc aagagttgac acattgtatt tccaaagtcc
     5281 cactgggcgg cagcatagtg agcccaccat agagcccacc gcatccccag catgcctgct
     5341 attgtcttcc caatcctccc ccttgctgtc ctgccccacc ccacccccca gaatagaatg
     5401 acacctactc agacaatgcg atgcaatttc ctcattttat taggaaagga cagtgggagt
     5461 ggcaccttcc agggtcaagg aaggcacggg ggaggggcaa acaacagatg gctgg
//
