Fig. 1.

The scheme for constructing improved degradome library. For sequencing purposes, the degradome library generated by this method can be treated as small RNA library and the resulting reads are ~ 27 nt long. The procedure includes: (1) poly(A) RNA isolation; (2) 5′RNA adapter ligation to uncapped poly(A) RNA with 5′ monophosphate; (3) reverse transcription to generate 1st strand cDNA using an oligo(dT)-tailed adapter (RT-primer); (4) second strand synthesis (1st PCR amplification); (5) EcoP15I digestion to generate ~ 27 nt long reads; (6) ligation of EcoP15I digestion products with a 3′ds-DNA adapter; (7) purification of ligation products on a PAGE gel; (8) degradome library enrichment (2nd PCR amplification); (9) purification of the final product on a PAGE gel; (10) library pooling and sequencing using Illumina HiSeq platform