Description
Hieff NGS™ MaxUp Human rRNA Depletion Kit (rRNA & ITS/ETS) is designed to remove rRNA and 45S ITS/ETS from human, mouse and rat total RNA based on RNase H-based workflow and to retain mRNA and other non-coding RNA. This kit is suitable for both intact and partially degraded RNA samples (e.g., FFPE RNA). Since degraded FFPE samples usually contain a higher proportion of ITS/ETS than fresh tissue samples, this kit adds probes in the Human 45S ITS/ETS region, and ITS removal can significantly increase the proportion of valid data in raw data.
Features
- Strong specificity: specifically remove rRNA and ITS/ETS from human samples, especially for FFPE samples
- High compatibility of template starting amount: applicable to 100 ng~1μg sample
- High removal effect: for rRNA, ITS and ETS in human\mouse and rat samples, the removal effect is more than 95%
- Stable quality: strict batch performance and stability quality control
Applications
- Gene Expression Research
- Alternative splicing analysis
- Detection and discovery of non coding RNA
- dentification of selective polyadenylation sites
- Fusion gene detection
Specifications
| Depletion Technology | RNase H |
| Sample Type | Total RNA of human, mouse and rat |
| Final Product Type | mRNA and other non coding RNAs |
| No. of Reactions | 24/96 Preps |
| Starting Material Amount | 100 ng~1 μg total RNA |
| Target | Remove rRNA and 45S ITS/ETS form human total RNA |
Components
| Components No. | Name | 12257ES24 (24T) | 12257ES96 (96T) |
| 12257-A | Hybridization Buffer | 72 μL | 288 μL |
| 12257-B | Probe Mix (rRNA & ITS/ETS) | 48 μL | 192 μL |
| 12257-C | RNase H Buffer | 72 μL | 288 μL |
| 12257-D | RNase H | 48 μL | 192 μL |
| 12257-E | DNase I Buffer | 660 μL | 2×1320 μL |
| 12257-F | DNase I | 60 μL | 240 μL |
Shipping and Storage
All the components are shipped with dry ice and can be stored at -15℃ ~ -25℃ for one year.
Figures
- rRNA removal specificity
For conventional samples, rRNA and ITS/EST can be removed by more than 98%, while for FFPE samples with integrity of about 50%, rRNA and ITS/EST can be removed by more than 95%. At the same time, FFPE with poor integrity can also be removed effectively.
Table 1. rRNA Removal specificity effect
| RNA quality | Removal scheme | Input | rRNA (%) | ITS/ETS (%) | Mapping (%) |
| 293T RNA; RIN=10 | remove rRNA | 1μg | 0.19 | 4.6 | 98.28 |
| remove rRNA and ITS/ETS | 1μg | 0.15 | 0.04 | 98.79 | |
| FFPE RNA; DV200 =90% | remove rRNA | 500ng | 0.23 | 4.75 | 95.35 |
| remove rRNA and ITS/ETS | 500ng | 0.21 | 0.02 | 95.42 | |
| FFPE RNA; DV200 =50% | remove rRNA | 500ng | 1.4 | 16.28 | 95.57 |
| remove rRNA and ITS/ETS | 500ng | 0.56 | 0.11 | 95.51 | |
| FFPE RNA; DV200 =20% | remove rRNA | 1μg | 0.35 | 67.61 | 58.4 |
| remove rRNA and ITS/ETS | 1μg | 0.79 | 0.84 | 60.35 |
Note: Different human RNA samples were used to remove rRNA or rRNA and ITS/ETS, and to construct libraries with RNA library Prep kit after ITS/ETS. The proportions of rRNA and ITS/ETS in offline data were analyzed by sequencing.
- Sequencing data performance
Table 2. Sequencing data performance
| Sample | DV200 | Kit | Clean Q20 (%) | Clean Q30 (%) | Clean GC (%) | rRNA (%) | ITS/ETS (%) | unique(%) |
| 1 | 24% | YEASEN Cat#12257 | 96.75 | 92.68 | 57.05 | 4.58 | 2.42 | 96.90 |
| 2 | 40% | 96.56 | 92.42 | 55.31 | 1.93 | 0.63 | 96.45 | |
| 3 | 53% | 97.4 | 93.52 | 45.19 | 0.23 | 0.01 | 98.08 | |
| 4 | 76% | 97.67 | 93.78 | 51.28 | 0.44 | 0.01 | 93.63 |
[1] Tian S, Zhang B, He Y, et al. CRISPR-iPAS: a novel dCAS13-based method for alternative polyadenylation interference. Nucleic Acids Res. 2022;50(5):e26. doi:10.1093/nar/gkac108(IF:16.971)