The initial template is prepared with 0.1 µg-1 µg of total RNA of human, mouse, or rat. Compared with traditional transcriptome library preparation kits, this kit enables the depletion of ribosomal RNA (rRNA), including both cytoplasmic (28S, 18S, 5.8S, and 5S) and mitochondrial (16S and 12S) rRNA, leaving mRNA and other non-coding RNA behind. This kit is suitable for subsequent non-coding RNA analysis such as lncRNA. Degraded RNA (i.e. FFPE RNA) could also be used to prepare the library with this kit. In addition, this kit enables the insertion of dUTP during the 2nd strand synthesis of cDNA. The double strand cDNA are digested by uracil-DNA glycosylase (UDG) to remove the second strand before sequencing. As a result, only information from the 1st strand cDNA is preserved. In addition to standard transcriptome information, strand-specific (i.e. from sense or anti-sense DNA) information can also be obtained from NGS data analysis.
Kit Components:
| Components | NR603-01 (24 rxn) | NR603-02 (96 rxn) | |
| NR 4 | rRNA Probe (H/M/R) | 24 μl | 96 μl |
| Probe Buffer | 72 μl | 288 μl | |
| RNaseH Buffer | 24 μl | 96 μl | |
| RNaseH | 96 μl | 384 μl | |
| DNase I Buffer | 672 μl | 4 × 672 μl | |
| DNase I | 48 μl | 192 μl | |
| NR 5 | Frag/Prime Buffer | 444 μl | 2 × 888 μl |
| Actinomycin D (5 mg/ml) | 24 μl | 96 μl | |
| 1st Strand Buffer | 144 μl | 576 μl | |
| 1st Strand Enzyme Mix | 48 μl | 192 μl | |
| 2nd Strand Marking Buffer | 480 μl | 2 × 960 μl | |
| 2nd Strand/End Repair Enzyme Mix | 120 μl | 480 μl | |
| NR 6 | dA-Tailing Buffer Mix | 240 μl | 960 μl |
| dA-Tailing Enzyme Mix | 60 μl | 240 μl | |
| Ligation Mix | 60 μl | 240 μl | |
| Stop Ligation Mix | 120 μl | 480 μl | |
| PCR Primer Mix | 120 μl | 480 μl | |
| Amplification Mix | 600 μl | 4 × 600 μl | |
| Heat-labile UDG | 24 μl | 96 μl |