Surface-enhanced Raman scattering (SERS) spectroscopy has been considered as a promising way to realize real-time, in-situ and ultrasensitive analysis of chemoand biochemical molecules in different applications even in intracellular or aqueous environments. In this work, polymer-supported novel Ag nano-necklaces (AgNLs) as flexible SERS substrates were fabricated for ultrasensitive chemical and biological detection. With the stringing of dense "hot spot" in three-dimension, AgNLs located on polydimethylsiloxane (PDMS) work like the removable and reusable "tip" on the surface of analytes with different morphologies and conditions. The novel substrate shows ultra-high enhancement factor (as high as 10 9 ) with excellent reproducibility and long-term stability (7 months) in an aqueous environment. With further functionalizing with p-mercaptobenzoic acid, AgNLs/PDMS elastomer also reveals sensitive and consistent pH detection ability over the wide range of pH 4.0-9.0, indicating their wide applications in biological and environmental fields. This work provides a feasible strategy for designing ultrasensitive, reproducible and flexible SERS substrate for practical detection.