LOCUS       sequence-373370-        4630 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  encodes zinc finger nuclease specific to AAVS1 locus (left).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4630)
  AUTHORS   Sim X, Cardenas-Diaz FL, French DL, Gadue P
  TITLE     A Doxycycline-Inducible System for Genetic Correction of iPSC 
            Disease Models.
  JOURNAL   Methods Mol Biol. 2015 Jan 29.
  PUBMED    25630922
REFERENCE   2  (bases 1 to 4630)
  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: "Methods Mol
            Biol. 2015 Jan 29."
COMMENT     Sequence Label: PGK-AAVS1ZFNL
FEATURES             Location/Qualifiers
     source          1..4630
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        406..905
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(441..464)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     838..857
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      932..941
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             944..964
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(2409..2425)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2409..2425)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2422..2444)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2433..2449
                     /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(2457..2463,2464..2481,2482..2487))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2502..2523
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2640..2657)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2811..3399)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2891..2910)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3570..4361,4362..4430))
                     /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     4193..4212
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4431..4535)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4603..4621
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt ctaccgggta ggggaggcgc
      421 ttttcccaag gcagtctgga gcatgcgctt tagcagcccc gctgggcact tggcgctaca
      481 caagtggcct ctggcctcgc acacattcca catccaccgg taggcgccaa ccggctccgt
      541 tctttggtgg ccccttcgcg ccaccttcta ctcctcccct agtcaggaag ttcccccccg
      601 ccccgcagct cgcgtcgtgc aggacgtgac aaatggaagt agcacgtctc actagtctcg
      661 tgcagatgga cagcaccgct gagcaatgga agcgggtagg cctttggggc agcggccaat
      721 agcagctttg ctccttcgct ttctgggctc agaggctggg aaggggtggg tccgggggcg
      781 ggctcagggg cgggctcagg ggcggggcgg gcgcccgaag gtcctccgga ggcccggcat
      841 tctgcacgct tcaaaagcgc acgtctgccg cgctgttctc ctcttcctca tctccgggcc
      901 tttcgacctg caagtcgacc cgcggaccgg tgccaccatg gctccaaaga aaaagcggaa
      961 agtgggaatc cacggagtcc ccgccgctat ggcagagaga ccttttcagt gccgaatttg
     1021 tatgcggaac ttctcttata attggcacct gcagaggcat atccgcactc acaccggaga
     1081 gaaacccttt gcatgcgaca tttgtggcag aaagttcgcc aggtccgatc atctgaccac
     1141 acacactaaa atccataccg gctctcagaa gccctttcag tgccgcattt gtatgcgaaa
     1201 cttctctcac aattacgccc gggactgcca catcagaaca catactgggg agaaaccttt
     1261 tgcttgcgat atttgtggac gaaagttcgc acagaacagt acacggatcg gccacactaa
     1321 aattcatctg agggggtctc agctggtgaa gagtgaactg gaggagaaga agagcgagct
     1381 gcgccacaaa ctgaagtacg tccctcatga gtatatcgaa ctgatcgaga ttgccaggaa
     1441 ttcaacccag gaccgcattc tggaaatgaa agtgatggag ttctttatga aggtctacgg
     1501 atatagaggc aaacacctgg gcgggtcaag gaagcctgat ggcgctatct acactgtggg
     1561 gagcccaatc gactatggag tgattgtcga taccaaggcc tacagcggag gctataacct
     1621 gcccattggc caggctgacg agatggaacg ctacgtggag gaaaaccaga cacgaaacaa
     1681 gcatctgaac ccaaatgaat ggtggaaggt gtatcccagc tccgtcactg agttcaagtt
     1741 tctgttcgtg agcgggcact ttaaaggaaa ctacaaggca cagctgaccc ggctgaatca
     1801 tatcacaaac tgtaatggcg ccgtgctgtc cgtcgaggaa ctgctgatcg ggggagaaat
     1861 gatcaaggcc ggcaccctga cactggagga agtgcggaga aagtttaaca atggggagat
     1921 caatttctga gcggccgctt gcagaaattg atgatctatt aaacaataaa gatgtccact
     1981 aaaatggaag tttttcctgt catactttgt taagaagggt gagaacagag tacctacatt
     2041 ttgaatggaa ggattggagc tacgggggtg ggggtggggt gggattagat aaatgcctgc
     2101 tctttactga aggctcttta ctattgcttt atgataatgt ttcatagttg gatatcataa
     2161 tttaaacaag caaaaccaaa ttaagggcca gctcattcct cccactcatg atctatagat
     2221 ctatagatct ctcgtgggat cattgttttt ctcttgattc ccactttgtg gttctaagta
     2281 ctgtggtttc caaatgtgtc agtttcatag cctgaagaac gagatcagca gcctctgttc
     2341 cacatacact tcattctcag tattgttttg ccaagttcta attccatcag aagcttggcg
     2401 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     2461 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     2521 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     2581 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     2641 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     2701 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     2761 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     2821 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     2881 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     2941 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3001 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3061 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3121 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3181 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3241 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3301 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3361 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3421 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3481 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3541 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3601 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3661 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3721 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3781 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     3841 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     3901 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     3961 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4021 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4081 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4141 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4201 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4261 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4321 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4381 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4441 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4501 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4561 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
     4621 ccctttcgtc
//
