New England Biolabs (NEB^®) nnounced the launch of the NEBNext Low-bias Small RNA Library Prep Kit, designed to minimize biased representation of small RNA species in sequencing data. This next generation small RNA preparation method is faster, less biased, and has a broader input range than other commercially available kits.

"Small RNA species – including rare and non-coding RNAs – have historically been relegated to a black box. Now, researchers are opening that box and realizing these tiny RNAs have an outsized influence on health and disease," said Betsy Young, Ph.D., Senior Product Marketing Manager for the NEBNext portfolio. "However, sequencing bias in preexisting library preparation methods has made it hard for researchers to understand the true diversity and abundance of small RNAs in their samples. This kit was designed to generate RNA libraries with a more accurate representation of small RNA species."

Protocol enhancements include the addition of a novel splint adaptor that increases the diversity of interactions, facilitating ligation and increasing sensitivity, with a streamlined, simplified protocol. As a result, researchers can now confidently analyze all RNA species present in biologically relevant samples. Additional improvements include unprecedented speed (~3.5 hrs), shelf life (18 months), and input range. Standard and 2´-O-methylated samples can also be processed using the same protocol, with multiplexing enabled through up to 480 compatible UDI primer pairs (available separately).

The kit's novel approach to adaptor ligation culminated from over a decade of research by NEB scientists. The first study, published in 2012, recognized that RNA sequencing bias was significant and caused mostly by unpredictable differences in ligation efficiency for small RNAs. The second foundational study, published in 2020, presented a novel library preparation workflow that reduced bias and increased sensitivity in small RNA library prep. Several protocol enhancements have since improved the kit to outperform the originally published methods.

"Bias in small RNA sequencing has long complicated the interpretation of RNA sequencing data, and limited RNA research," explained Greg Lohman, Research Group Leader in NEB's Molecular Enzymology Division. "We anticipate that successfully resolving this bias through a novel library preparation method will make diverse RNA research easier and open the door to future discoveries."