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Table 2 The algorithm for small RNA processing for the E. histolytica Rahman size-fractionated small RNA library

From: Small RNA pyrosequencing in the protozoan parasite Entamoeba histolytica reveals strain-specific small RNAs that target virulence genes

Procedures

Output

5-P independent cloning (5CIP+PNK method)

Rahman total RNA, size fractionation (15-30nt) library

Total reads by Pyrosequencing

151,656 reads

Primer trimming, size-limiting (15–40nt)

140,571 reads

Unique sequences (percentage)

98,414 unique sequences (82,780 cloned only once; 84.1%) (15,634 cloned more than once; 15.9%)

Scan for tRNA, rRNA

tRNA: 517 sequences rRNA:1,753 sequences

Scan for SINE/LINE, EhERE elements

4,439 sequences

Map to E. histolytica genome

52,028 sequences

Map to E. histolytica predicted ORFs

35,157 sequences

  1. Half of a pyrophosphate sequencing run was performed. Unix tools were used to remove adaptor and linker sequences. The Bowtie alignment tool was used to scan for structural RNAs, repetitive elements and for the final genome mapping. We used E. histolytica HM-1:IMSS genome as a reference genome due to the E. histolytica Rahman sequence not being assembled. The numbers of reads that map to the genome sequence at each step are listed.