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Table 1 Mutagenic oligos (Round 1 PCR) used for site directed mutagenesis in exon 8 and 9 of the WT1 gene

From: Screening of WT1 mutations in exon 8 and 9 in children with steroid resistant nephrotic syndrome from a single centre and establishment of a rapid screening assay using high-resolution melting analysis in a clinical setting

Mutation

Oligo Name

Oligo sequence 5’–3’

Annealing temperature (°C)

Exon 8

c.1079G > A;C360Y (hg19 - c.1283G > A; C428Y)

WT1_E8_F _A

E8_C428Y_R_C

TTCCCCAAGGTGAGAAACCA

ACCTTCGTTCATAGTCCTTGAAG

56

E8_C428Y_F_ B

WT1_E8_R_D

CTTCAAGGACTATGAACGAAGGT

GCTGCCAGCAATGAGAAGTG

58

Exon 8

c.1119C > A;H373Q (hg19 - c.1323C > A;H441Q)

WT1_E8_F _A

E8_H441Q_R_C

TTCCCCAAGGTGAGAAACCA

TGTCTCCTTTGTTGTCTTTTGAG

58

E8_ H441Q _F_ B

WT1_E8_R_D

CTCAAAAGACAACAAAGGAGACA

GCTGCCAGCAATGAGAAGTG

58

Exon 9

c.1180C > T;R394W (hg19 - c.1384C > T;R462W)

WT1_E9_F_A

E9_R462W_R_C

TTGTTAGGGCCGAGGCTAGA

TGGTCGGACCAGGAGAAC

56

E9_R462W_F _B

WT1_E9_R _D

GTTCTCCTGGTCCGACCA

AGTGCGTAAACTTTTCTTCACAT

60

Exon 9

c.1190A > C;H397P (hg19 - c.1394A > C; H465P)

WT1_E9_F_A

E9_ H465P_R_C

TTGTTAGGGCCGAGGCTAGA

GTCTTCAGGGGGTCGGACC

60

E9_ H465_F_B

WT1_E9_R _D

GGTCCGACCCCCTGAAGAC

AGTGCGTAAACTTTTCTTCACAT

52

Exon 9

c.A1200C > T;H401Y (hg19 - c.1405C > T;H469Y)

WT1_E9_F_A

E9_ H469Y_R_C

TTGTTAGGGCCGAGGCTAGA

GTCCTGGTGTGAGTCTTCAGGTG

58

E9_ H469Y_F_B

WT1_E9_R _D

CACCTGAAGACTCACACCAGGAC

AGTGCGTAAACTTTTCTTCACAT

56

  1. Oligos in bold indicate mutagenic oligos