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Development of a mutation hotspot detection kit for the phenylalanine hydroxylase gene by ARMS-PCR combined with fluorescent probe technology
  • +12
  • jinhua he,
  • Rong Qiang,
  • Lin Wang,
  • weijie xu,
  • wei li,
  • Na Cai,
  • Xiao Bin Wang,
  • RuiXue Zhang ,
  • Li Ping Zhang,
  • Xiao Ping Ma,
  • Chen Wei ,
  • ChengRong Song,
  • WenWen Yu,
  • Xiang Wang,
  • Xu LI
jinhua he
pan yu central hospital
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Rong Qiang
Northwest Women and Children's Hospital
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Lin Wang
Medical heredity research center, northwest Women’s and children’s hospital, shaanxi,Xi'an, 710003,People’s republic of china
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weijie xu
Guangzhou Lanji Biotechnology Co., Ltd., Guangzhou, Guangdong 510665, P. R. China
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wei li
Medical heredity research center, northwest Women’s and children’s hospital, shaanxi,Xi'an, 710003,People’s republic of china
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Na Cai
Medical heredity research center, northwest Women’s and children’s hospital, shaanxi,Xi'an, 710003,People’s republic of china
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Xiao Bin Wang
Medical heredity research center, northwest Women’s and children’s hospital, shaanxi,Xi'an, 710003,People’s republic of china
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RuiXue Zhang
Medical heredity research center, northwest Women’s and children’s hospital, shaanxi,Xi'an, 710003,People’s republic of china
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Li Ping Zhang
Medical heredity research center, northwest Women’s and children’s hospital, shaanxi,Xi'an, 710003,People’s republic of china
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Xiao Ping Ma
Medical heredity research center, northwest Women’s and children’s hospital, shaanxi,Xi'an, 710003,People’s republic of china
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Chen Wei
Medical heredity research center, northwest Women’s and children’s hospital, shaanxi,Xi'an, 710003,People’s republic of china
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ChengRong Song
Medical heredity research center, northwest Women’s and children’s hospital, shaanxi,Xi'an, 710003,People’s republic of china
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WenWen Yu
"Center for Translational Medicine, the First Affiliated Hospital,Xi'an Jiaotong University;Xi'an,710061,P.R.China. Key Laboratory for Tumor precision Medicine of Shaanxi Province. the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, 710061, P.R. China."
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Xiang Wang
"Center for Translational Medicine, the First Affiliated Hospital,Xi'an Jiaotong University;Xi'an,710061,P.R.China. Key Laboratory for Tumor precision Medicine of Shaanxi Province. the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, 710061, P.R. China."
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Xu LI
"Center for Translational Medicine, the First Affiliated Hospital,Xi'an Jiaotong University;Xi'an,710061,P.R.China. Key Laboratory for Tumor precision Medicine of Shaanxi Province. the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, 710061, P.R. China. "
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Abstract

To develop a screening kit for detecting mutation hotspots of the PAH gene.. We designed and developed a screening kit to detect 9 mutation sites covering more than 50% of the PAH mutations found in Shaanxi province (c.728G>A, c.1197A>T, c.331C>T, c.1068C>A, c.611A>G, c.1238G>C, c.721C>T, c.442-1G>A, and c.158G>A) by using amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) combined with fluorescent probe technology. Peripheral blood and dried blood samples from PKU families were used for clinical verification of the newly developed kit. PAH gene mutations were detected in 84 children diagnosed with PKU. A total of 159 mutant alleles were identified, consisting of 100 missense mutations, 28 shear mutations, 24 nonsense mutations, and 7 deletion mutations. Exon 7 had the highest mutation frequency (32.08%).the mutation frequency of p.R243Q was the highest, accounting for 20.13% of all mutations, followed by p.R111X, IVS4-1G>A, EX6-96A>G, and p.R413P; these 5 loci accounted for 47.17% (75/159) of all mutations. Fifteen mutation sites were identified in the 47 PAH carriers identified by NCS, which were verified by the newly developed kit, with an agreement rate of 100%.This newly developed kit based on ARMS-PCR combined with fluorescent probe technology can be used to detect common PAH gene mutations.