Effect of Spacer Length on Capturing Performance of Multivalent Aptamers Generated by Rolling Circle Amplification

Title: Effect of Spacer Length on Capturing Performance of Multivalent Aptamers Generated by Rolling Circle Amplification
Authors: Wang, Zhong
Date: 2022-06-21
Abstract: Multivalent aptamer refers to a technique that joins two or more identical or different types of aptamer monomers together, with or without the presence of structural or other functional elements. As a rapid and easy method for fabricating the multivalent aptamer constructs, rolling circle amplification (RCA) has attracted great attention in recent decades. The incorporation of properly designed structural elements, such as intra-molecular spacers, have been shown to greatly enhance the efficiency of the multivalent aptamer system [1]. The objective of this current study is to systemically investigate the effect of different lengths of poly thymine spacer designs (polyT, from no spacer/NT, 5T, 10T, and 15T) on the capturing performance of RCA-generated multivalent system. To achieve this, we designed four circular probe templates by inserting zero, five, ten, and fifteen adenine bases (polyA). These polyA domains in the circular probe template are complementary to polyT with respective lengths in between the adjacent aptamers on the resultant RCA products (RCAPs). We found that the resultant RCAPs with length shorter than 10T showed a lack or low ability to capture target cells E.Coli O157:H7. When spacer lengths reach or exceed 10T, the capturing performance of the respective multivalent aptamer chain is significantly enhanced. This phenomenon can be explained by larger hydrodynamic sizes and less nonspecific secondary structures observed in RCAP with spacer length no less than 10T. Moreover, we found that there is also a trade-off that the number of polyA bases added into the circular probe template can significantly impair the efficiency of RCA reaction in respective to cyclization yield and amplification rate. The results of this research explain with details how the design of spacer affects RCA reaction efficiency and RCAPs’ capturing performance, which provides ideas in designing an efficient RCA-generated multivalent system.
URL: http://hdl.handle.net/10393/43720
CollectionThèses, 2011 - // Theses, 2011 -