Depletion probes

A method of depleting a target RNA transcript in a sample, the method comprising contacting the sample with:

• a nuclease that recognizes DNA/RNA heteroduplexes and cleaves the RNA;
• and
• a plurality of DNA oligos that are each complementary to the target RNA transcript and have minimal to no annealing to off-target transcripts, wherein:
• the sequences of the DNA oligos of the plurality of DNA oligos are selected by:
• (i) generating a set of candidate sequences complementary to the target RNA transcript, wherein the candidate sequences have a predicted melting temperature within a 3.0 degree variance around an initial value;
• (ii) removing from the set of candidate sequences, candidate sequences that do not have minimal to no annealing to off-target transcripts;
• and
• (iii) selecting from the set of candidate sequences, sequences of the plurality of DNA oligos using a cost function to minimize gaps between adjacent DNA/RNA heteroduplexes formed by the target RNA transcript and the plurality of DNA oligos,
• wherein the DNA oligos of the plurality of DNA oligos are present in the sample at non-equimolar concentrations;
• wherein the plurality of DNA oligos form DNA/RNA heteroduplexes with the target RNA transcript in the sample and gaps between the DNA/RNA heteroduplexes are 0-111 nucleotides.