LOCUS       sequence-319289-        3504 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used as a donor vector for the targeted correction of a mutation in 
            PIGA exon 6.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3504)
  AUTHORS   Hyodo T, Rahman ML, Karnan S, Ito T, Toyoda A, Ota A, Wahiduzzaman 
            M, Tsuzuki S, Okada Y, Hosokawa Y, Konishi H
  TITLE     Tandem Paired Nicking Promotes Precise Genome Editing with Scarce 
            Interference by p53.
  JOURNAL   Cell Rep. 2020 Jan 28;30(4):1195-1207.e7. doi: 
            10.1016/j.celrep.2019.12.064.
  PUBMED    31995758
REFERENCE   2  (bases 1 to 3504)
  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"; doi: 
            "10.1016/j.celrep.2019.12"; journalName: "Cell Rep"; date: 
            "2020-01-28- 28"; volume: "30"; issue: "4"; pages: "1195-1207.e7"
COMMENT     Sequence Label: Donor-PIGAshort
FEATURES             Location/Qualifiers
     source          1..3504
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1316..1336)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1316..1334)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1355..1371)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1355..1371)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1368..1390)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1379..1395
                     /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(1403..1409,1410..1427,1428..1433))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1448..1469
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1586..1603)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1757..2345)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1837..1856)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2516..3307,3308..3376))
                     /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     3139..3158
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3377..3481)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggagctcca
      661 ccgcggtggc ggccgctcta gactttgcgg gactttggaa ttctagtagt tggctaaaat
      721 tgatggtgac tccctagggg agaggaactg tggcttattc tttgtatcac acagcctcta
      781 gcccggtgtc ttgcccagaa taagtaccca ataagttttg aatcaaatgg caataattat
      841 cgtgattaag ttcattaagg atatcattac tatgacaaca acaacaaaac acaagcaact
      901 tttgtttgta tttgtttata aaatgttcct gcaggatcgg gtcattgttt atcatgggac
      961 aggtgatggg gtgatgggtg aaagtgctca cacttggctt ctctccctcc aggtatatga
     1021 ccgggtatca gtggaagctg tgttgccaat ggacaaacga ctggacagac ttatttctca
     1081 ctgcggccca gtaacaggct acatctttgc tttgttggca gttttcaact tcctcttcct
     1141 cattttcttg agatggatga ctccagattc tatcattgat gttgcaatag atgccactgg
     1201 gccacggggt gcctggacta ataactattc tcacagtaaa agagggggtg agaataatga
     1261 gatatctgaa accaggtaga agctcgaggg ggggcccggt acccagcttt tgttcccttt
     1321 agtgagggtt aattgcgcgc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     1381 gttatccgct cacaattcca cacaacatac gagccgggag cataaagtgt aaagcctggg
     1441 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     1501 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     1561 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     1621 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     1681 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     1741 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     1801 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     1861 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     1921 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     1981 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     2041 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     2101 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     2161 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     2221 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     2281 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     2341 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     2401 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     2461 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     2521 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     2581 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     2641 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     2701 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     2761 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     2821 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     2881 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     2941 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     3001 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     3061 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     3121 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     3181 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     3241 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     3301 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     3361 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     3421 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     3481 gcacatttcc ccgaaaagtg ccac
//
