19 nt RNA size marker	5′CGUACGCGGGUUUAAACGA
35 nt RNA size marker	5′ CUCAUCUUGGUCGUACGCGGAAUAGUUUAAACUGU
5′ adapter	GUUCAGAGUUCUACAGUCCGACGAUC
DNA oligonucleotides
3′ PCR primer	CAAGCAGAAGACGGCATACGA
5′ PCR primer	AATGATACGGCGACCACCGACAGGTTCAGAGTTCTACAGTCCGA
3′ adapters*
Ad01	App-TCACTTCGTATGCCGTCTTCTGCTTG-L
Ad02	App-TCATCTCGTATGCCGTCTTCTGCTTG-L
Ad03	App-TCCACTCGTATGCCGTCTTCTGCTTG-L
Ad04	App-TCCGTTCGTATGCCGTCTTCTGCTTG-L
Ad05	App-TCCTATCGTATGCCGTCTTCTGCTTG-L
Ad06	App-TCGATTCGTATGCCGTCTTCTGCTTG-L
Ad07	App-TCGCGTCGTATGCCGTCTTCTGCTTG-L
Ad08	App-TCTAGTCGTATGCCGTCTTCTGCTTG-L
Ad09	App-TCTCCTCGTATGCCGTCTTCTGCTTG-L
Ad10	App-TCTGATCGTATGCCGTCTTCTGCTTG-L

^*Listed are ten 3′ adapters we frequently use in the lab [18,19]. App, 5′ terminal adenosine residue connected via a 5′,5′-diphosphate bridge to the 5′OH of the 5′ nucleotide; L, 3′ aminohexyl blocking group. Oligonucleotides can be obtained from suppliers of customized oligonucleotides [e.g. IDT (http://www.idtdna.com) or MultiplexDX (http://www.multiplexdx.com)]. Note: to further reduce possible biases in ligation efficiency randomized nucleotides at the 5′ end of the 3′ adapter, as well as on the 3′ end of the 5′ adapter can be introduced. Nevertheless, we do not find substantial improvement in library representation by these modifications, consistent with our previous finding that secondary structure formation of RNA substrates contributed most to ligation biases [17].