> For the complete documentation index, see [llms.txt](https://knowledge.illumina.com/llms.txt). Markdown versions of documentation pages are available by appending `.md` to page URLs; this page is available as [Markdown](https://knowledge.illumina.com/library-preparation/rna-library-prep/library-preparation-rna-library-prep-faq-list/000001397.md).

# TruSeq Small RNA input best practices

**Best Practices**

* This kit is designed to start with total RNA (although pre-purified miRNA can be used). It is critical that this total RNA be high-quality, with a RIN of at least 8. Any standard RNA isolation technique can be used, as long as it retains miRNA (for example, some spin column RNA purifications will not capture the miRNA fraction). Use of mildly degraded RNA will result in low yields and loss of the low end of the dynamic range of the assay. More severely degraded total RNA will most likely produce low or no yield.
* The total RNA is suspended in nuclease-free water or 10 mM Tris. DEPC will not interfere with the enzymes in this kit.
* This kit is designed for use with 1 µg of total RNA. Illumina does not support starting material amounts lower than 1 µg.

**What is the minimum input material?**\
The protocol is optimized using 1.0 µg of total RNA.

**Can this protocol use purified small RNA as starting material?**\
Yes. Note that there can be high sample variability using some commercially available small RNA spin column purification methods, and validation of any purification method may be necessary. If purified small RNA is used as starting material, 10-50 ng should be used.

**Does the TruSeq small RNA protocol work with plant small RNA?**\
Some plant small RNA molecules are methylated at their 3' end. The RNA ligase used is somewhat able to ligate to several common methyl modifications with sufficient efficiency to support library construction, though efficiency may be lower.

\
\
\ <br>

|                                                                                                                                                                                                                                                                                                                                                                              |
| :--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------: |
| *For any feedback or questions regarding this article (Illumina Knowledge Article #1397), contact Illumina Technical Support* [*techsupport@illumina.com*](mailto:techsupport@illumina.com?subject=Question%2FFeedback%20Regarding%20Illumina%20Knowledge%20Article%20#000001397%20-%20Library%20Preparation%20\&body=Dear%20Illumina%20Technical%20Support,%0D%0A%0D%0A)*.* |


---

# Agent Instructions
This documentation is published with GitBook. GitBook is the documentation platform designed so that both humans and AI agents can read, navigate, and reason over technical content effectively. Learn more at gitbook.com.

## Querying This Documentation
If you need additional information that is not directly available in this page, you can query the documentation dynamically by asking a question.

Perform an HTTP GET request on the current page URL with the `ask` query parameter, and the optional `goal` query parameter:

```
GET https://knowledge.illumina.com/library-preparation/rna-library-prep/library-preparation-rna-library-prep-faq-list/000001397.md?ask=<question>&goal=<endgoal>
```

`ask` is the immediate question: it should be specific, self-contained, and written in natural language.
`goal` is optional and describes the broader end goal you are ultimately trying to accomplish on behalf of the user. GitBook uses it to tailor the answer towards what is most useful for that goal.

The response will contain a direct answer to the question and relevant excerpts and sources from the documentation.

Use this mechanism when the answer is not explicitly present in the current page, you need clarification or additional context, or you want to retrieve related documentation sections.