Taqman-MGB nanoPCR for Highly Specific Detection of Single-Base Mutations

Zhenrui Xue,1– 3,* Minli You,2,3,* Ping Peng,1– 3,* Haoyang Tong,2,3 Wanghong He,2– 4 Ang Li,4 Ping Mao,1– 3 Ting Xu,1 Feng Xu,2,3 Chunyan Yao1 1Department of Transfusion Medicine, Southwest Hospital, Third Military Medical University, Army Medical University,...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Xue Z, You M, Peng P, Tong H, He W, Li A, Mao P, Xu T, Xu F, Yao C
Formato:	article
Lenguaje:	EN
Publicado:	Dove Medical Press 2021
Materias:	nanoparticle-assisted pcr point mutation gold nanoparticle specificity Medicine (General) R5-920
Acceso en línea:	https://doaj.org/article/dc51d91d62784257a7c5a5bada7fa7a6
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:dc51d91d62784257a7c5a5bada7fa7a6
record_format	dspace
spelling	oai:doaj.org-article:dc51d91d62784257a7c5a5bada7fa7a62021-12-02T15:02:03ZTaqman-MGB nanoPCR for Highly Specific Detection of Single-Base Mutations1178-2013https://doaj.org/article/dc51d91d62784257a7c5a5bada7fa7a62021-05-01T00:00:00Zhttps://www.dovepress.com/taqman-mgb-nanopcr-for-highly-specific-detection-of-single-base-mutati-peer-reviewed-fulltext-article-IJNhttps://doaj.org/toc/1178-2013Zhenrui Xue,1– 3,* Minli You,2,3,* Ping Peng,1– 3,* Haoyang Tong,2,3 Wanghong He,2– 4 Ang Li,4 Ping Mao,1– 3 Ting Xu,1 Feng Xu,2,3 Chunyan Yao1 1Department of Transfusion Medicine, Southwest Hospital, Third Military Medical University, Army Medical University, Chongqing, 400038, People’s Republic of China; 2The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China; 3Bioinspired Engineering and Biomechanics Center (BEBC), Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China; 4Key Laboratory of Shaanxi Province for Craniofacial Precision Medicine Research, College of Stomatology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China*These authors contributed equally to this workCorrespondence: Chunyan Yao; Feng XuDepartment of Transfusion Medicine, Southwest Hospital, Third Military Medical University, Army Medical University, Chongqing, 400038, People’s Republic of ChinaTel +64 389 619 2922Email yao_yao24@yahoo.com; fengxu@mail.xjtu.edu.cnPurpose: Detection of single-base mutations is important for real-time monitoring of tumor progression, therapeutic effects, and drug resistance. However, the specific detection of single-base mutations from excessive wild-type background sequences with routine PCR technology remains challenging. Our objective is to develop a simple and highly specific qPCR-based single-base mutation detection method.Methods: Using EGRF T790M as a model, gold nanoparticles at different concentrations were separately added into the Taqman-MGB qPCR system to test specificity improvement, leading to the development of the optimal Taqman-MGB nanoPCR system. Then, these optimal conditions were used to test the range of improvement in the specificity of mutant-type and wild-type templates and the detection limit of mutation abundances in a spiked sample.Results: The Taqman-MGB nanoPCR was established based on the traditional qPCR, with significantly suppressed background noise and improved specificity for single-base mutation detection. With EGFR T790M as a template, we demonstrated that our Taqman-MGB nanoPCR system could improve specificity across a wide concentration range from 10− 9 μM to 10 μM and detect as low as 0.95% mutation abundance in spiked samples, which is lower than what the traditional Taqman-MGB qPCR and existing PCR methods can detect. Moreover, we also proposed an experimentally validated barrier hypothesis for the mechanism of improved specificity.Conclusion: The developed Taqman-MGB nanoPCR system could be a powerful tool for clinical single-base mutation detection.Keywords: nanoparticle-assisted PCR, point mutation, gold nanoparticle, specificityXue ZYou MPeng PTong HHe WLi AMao PXu TXu FYao CDove Medical Pressarticlenanoparticle-assisted pcrpoint mutationgold nanoparticlespecificityMedicine (General)R5-920ENInternational Journal of Nanomedicine, Vol Volume 16, Pp 3695-3705 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	nanoparticle-assisted pcr point mutation gold nanoparticle specificity Medicine (General) R5-920
spellingShingle	nanoparticle-assisted pcr point mutation gold nanoparticle specificity Medicine (General) R5-920 Xue Z You M Peng P Tong H He W Li A Mao P Xu T Xu F Yao C Taqman-MGB nanoPCR for Highly Specific Detection of Single-Base Mutations
description	Zhenrui Xue,1– 3,* Minli You,2,3,* Ping Peng,1– 3,* Haoyang Tong,2,3 Wanghong He,2– 4 Ang Li,4 Ping Mao,1– 3 Ting Xu,1 Feng Xu,2,3 Chunyan Yao1 1Department of Transfusion Medicine, Southwest Hospital, Third Military Medical University, Army Medical University, Chongqing, 400038, People’s Republic of China; 2The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China; 3Bioinspired Engineering and Biomechanics Center (BEBC), Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China; 4Key Laboratory of Shaanxi Province for Craniofacial Precision Medicine Research, College of Stomatology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China*These authors contributed equally to this workCorrespondence: Chunyan Yao; Feng XuDepartment of Transfusion Medicine, Southwest Hospital, Third Military Medical University, Army Medical University, Chongqing, 400038, People’s Republic of ChinaTel +64 389 619 2922Email yao_yao24@yahoo.com; fengxu@mail.xjtu.edu.cnPurpose: Detection of single-base mutations is important for real-time monitoring of tumor progression, therapeutic effects, and drug resistance. However, the specific detection of single-base mutations from excessive wild-type background sequences with routine PCR technology remains challenging. Our objective is to develop a simple and highly specific qPCR-based single-base mutation detection method.Methods: Using EGRF T790M as a model, gold nanoparticles at different concentrations were separately added into the Taqman-MGB qPCR system to test specificity improvement, leading to the development of the optimal Taqman-MGB nanoPCR system. Then, these optimal conditions were used to test the range of improvement in the specificity of mutant-type and wild-type templates and the detection limit of mutation abundances in a spiked sample.Results: The Taqman-MGB nanoPCR was established based on the traditional qPCR, with significantly suppressed background noise and improved specificity for single-base mutation detection. With EGFR T790M as a template, we demonstrated that our Taqman-MGB nanoPCR system could improve specificity across a wide concentration range from 10− 9 μM to 10 μM and detect as low as 0.95% mutation abundance in spiked samples, which is lower than what the traditional Taqman-MGB qPCR and existing PCR methods can detect. Moreover, we also proposed an experimentally validated barrier hypothesis for the mechanism of improved specificity.Conclusion: The developed Taqman-MGB nanoPCR system could be a powerful tool for clinical single-base mutation detection.Keywords: nanoparticle-assisted PCR, point mutation, gold nanoparticle, specificity
format	article
author	Xue Z You M Peng P Tong H He W Li A Mao P Xu T Xu F Yao C
author_facet	Xue Z You M Peng P Tong H He W Li A Mao P Xu T Xu F Yao C
author_sort	Xue Z
title	Taqman-MGB nanoPCR for Highly Specific Detection of Single-Base Mutations
title_short	Taqman-MGB nanoPCR for Highly Specific Detection of Single-Base Mutations
title_full	Taqman-MGB nanoPCR for Highly Specific Detection of Single-Base Mutations
title_fullStr	Taqman-MGB nanoPCR for Highly Specific Detection of Single-Base Mutations
title_full_unstemmed	Taqman-MGB nanoPCR for Highly Specific Detection of Single-Base Mutations
title_sort	taqman-mgb nanopcr for highly specific detection of single-base mutations
publisher	Dove Medical Press
publishDate	2021
url	https://doaj.org/article/dc51d91d62784257a7c5a5bada7fa7a6
work_keys_str_mv	AT xuez taqmanmgbnanopcrforhighlyspecificdetectionofsinglebasemutations AT youm taqmanmgbnanopcrforhighlyspecificdetectionofsinglebasemutations AT pengp taqmanmgbnanopcrforhighlyspecificdetectionofsinglebasemutations AT tongh taqmanmgbnanopcrforhighlyspecificdetectionofsinglebasemutations AT hew taqmanmgbnanopcrforhighlyspecificdetectionofsinglebasemutations AT lia taqmanmgbnanopcrforhighlyspecificdetectionofsinglebasemutations AT maop taqmanmgbnanopcrforhighlyspecificdetectionofsinglebasemutations AT xut taqmanmgbnanopcrforhighlyspecificdetectionofsinglebasemutations AT xuf taqmanmgbnanopcrforhighlyspecificdetectionofsinglebasemutations AT yaoc taqmanmgbnanopcrforhighlyspecificdetectionofsinglebasemutations
_version_	1718389169052450816

Taqman-MGB nanoPCR for Highly Specific Detection of Single-Base Mutations

Ejemplares similares