Highly Precise and Sensitive Polymerase Chain Reaction Using Mesoporous Silica-Immobilized Enzymes.

Content Provider	Europe PMC
Author	Matsuura, Shun-ichi Baba, Tomoya Ikeda, Takuji Yamamoto, Katsutoshi Tsunoda, Tatsuo Yamaguchi, Aritomo
Copyright Year	2022
Abstract	A highly preciseand sensitive technology that enables DNA amplification/detectionfrom minimal amounts of nucleic acid is expected to find applicabilityin genetic testing involving small amounts of samples. The use ofa free enzyme in conventional DNA amplification techniques, such asthe polymerase chain reaction (PCR), frequently causes side reactions(i.e., nonspecific DNA amplification) when ≤103 substrateDNA molecules are present, thereby preventing selective amplificationof the target DNA. To address this issue, we have developed a novelDNA amplification system, mesoporous silica-enhanced PCR (MSE-PCR),which involves the immobilization of a thermostable DNA polymerasefrom Thermococcus kodakaraensis (KODDNA polymerase) into highly ordered nanopores of the mesoporous silicato control the reaction environment around the enzyme. In the MSE-PCRsystem using immobilized KOD DNA polymerase, such nonspecific DNAamplification was remarkably inhibited under the same conditions.Furthermore, the optimization of mesoporous silica pore sizes enabledselective and efficient DNA amplification from DNA substrates at thesingle-molecule level, i.e., one ten-thousandth of the amount of substrateDNA required for a DNA amplification reaction using a free enzyme.The results obtained in this study have shown that the nanopores ofmesoporous silica can inhibit nonspecific reactions in DNA amplification,thereby considerably improving the specificity and sensitivity ofthe DNA polymerase reaction.
ISSN	19448244
Journal	ACS Applied Materials & Interfaces
Volume Number	14
PubMed Central reference number	PMC9266630
Issue Number	26
PubMed reference number	35700238
e-ISSN	19448252
DOI	10.1021/acsami.2c01992
Language	English
Publisher	American Chemical Society
Publisher Date	2022-06-14
Access Restriction	Open
Rights License	Permits non-commercial access and re-use, provided that author attribution and integrity are maintained; but does not permit creation of adaptations or other derivative works (https://creativecommons.org/licenses/by-nc-nd/4.0/). © 2022 The Authors. Published by American Chemical Society
Subject Keyword	mesoporous silica materials polymerase chainreaction DNA polymerase immobilized enzyme DNA amplification
Content Type	Text
Resource Type	Article
Subject	Nanoscience and Nanotechnology Medicine Materials Science