RiboMinus™ 转录组分离试剂盒（人/小鼠）

纯化后的RNA可以方便的用260 nm 紫外吸光值或Quant-iT RNA Assay试剂盒进行定量。要验证rRNA的去除效果，可以对样本进行琼脂糖凝胶电泳检测或使用Agilent 2100 Bioanalyzer进行分析。与对照相比，样本的琼脂糖凝胶电泳应无18S和28S rRNA条带。通过琼脂糖凝胶电泳也可以判断样品是否有DNA污染及RNA降解。rRNA的去除效率，RNA的降解，以及RNA的浓度也都可以通过Bioanalyzer进行分析。

对于物种特异性来说，以下物种对于LSU/SSU探针没有任何错配(LSU (28S) / SSU (18S)探针完美配对）。这意味着探针可从下列物种中去除18S和28S序列。

陆生物种：Human, mouse, rat, frog, rabbit, cow, pig, chicken, Drosophila,* Caenorhabditis elegans, midge, mosquito, yeast (Saccharomyces cerevisiae, Schizosaccharomyces pombe)
海洋物种：Sea squirt, coelacanth, lancelet, eel, salmon, sturgeon, ratfish, lamprey, brown plankton, zooplankton
植物：Arabidopsis thaliana, Brassica napus, rice, tomato, Humulus lupulus, Zea mays, wheat, soybean, pine, aspen

* 该试剂盒中的28S和18S探针分别和果蝇的 18S和28S rRNA序列完全匹配。但是，有报道表明果蝇(昆虫)的 28S rRNA会分裂为2个较小片段并与18S共迁移。目前准确的分裂位点还未被找到。但是，基于用户的反馈，该试剂盒并不能完全去除分裂的28S rRNA。我们也没有用该试剂盒测试过果蝇样本。

5S和5.8S序列更具多样性，因此效果好的探针所使用的物种范围可能会较小：
5S： 0至1个错配：人，小鼠，大鼠，青蛙，鸡，果蝇，线虫， 拟南芥，玉米
5.8S： 0错配：人, 小鼠，大鼠

This is most likely due to contamination of the total RNA sample with genomic DNA. Before the RiboMinus Kit procedure, we recommend treating the total RNA sample with DNase I to remove any genomic DNA contamination.

The purified RNA is easily quantitated using UV absorbance at 260 nm or a Quant-iT RNA Assay Kit. To verify rRNA depletion, electrophorese your sample on an agarose gel or use an Agilent 2100 Bioanalyzer. Agarose gel electrophoresis should show depletion of 18S and 28S rRNA bands compared to a control sample. Absence of contaminating DNA and RNA degradation may also be confirmed by agarose gel electrophoresis. The efficiency of rRNA depletion in the purified RNA, RNA degradation, and RNA concentration can also be analyzed using a bioanalyzer.

With regard to species specificity, the following organisms have 0 mismatches for the LSU/SSU probes (LSU (28S) / SSU (18S) Probe perfect matches). This means the probes will remove the 18S and 28S sequences from the following species.

- Species from the Land: Human, mouse, rat, frog, rabbit, cow, pig, chicken, Drosophila,* Caenorhabditis elegans, midge, mosquito, yeast (Saccharomyces cerevisiae, Schizosaccharomyces pombe)
- Species from the Sea: Sea squirt, coelacanth, lancelet, eel, salmon, sturgeon, ratfish, lamprey, brown plankton, zooplankton
- Plants: Arabidopsis thaliana, Brassica napus, rice, tomato, Humulus lupulus, Zea mays, wheat, soybean, pine, aspen

* The 28S and 18S probes in this kit respectively align 100% to the Drosophila 18S and 28S rRNA sequence. However, there are reports suggesting Drosophila (insect) 28S rRNA split into 2 smaller fragments co-migrating with 18S. The exact split site is not identified yet. But based on customer feedback, this kit does not remove the split 28S rRNA completely. We have not tested Drosophila samples with this kit.

The 5S and 5.8S sequences are more divergent, and thus the best probes possible have reduced species breadth:
- 5S: zero to 1 mismatch: human, mouse, rat, frog, chicken, Drosophila, C. elegans, A. thaliana, Z. mays
- 5.8S: zero mismatches: human, mouse, rat