General guidelines regarding amplicon design
We recommend customers to design 1 amplicon per target (either on-target or off-target) and to avoid cases where there are either multiple targets in the same amplicon or multiple amplicons covering a target. This might not always be possible due to target coordinates as well as amplicon design constraints.
- If you are concerned about your on-target or off-target size/capture, please contact our support team (email@example.com).
- Note: The GE pipeline cannot analyze a target spanning more than 1 amplicon.
What size indel does the GE pipeline work with?
Our current analysis pipeline has been tested to capture INDELs up to 50 bp in length, if you observe larger INDELs in the final report please interpret those with caution. Additionally if you want to design a panel to cover and analyze larger deletions please contact the support team to design a panel optimized for larger targets.
What is the recommended panel size?
The recommended size for this product is 20-100 amplicons. If you need a larger panel, please contact support.
How much padding is recommended around on- and off- target sites?
This can vary based on the expected indel length and type of editing. If you are expecting a larger indel, we would recommend more padding up and downstream of your target.Please use the following example table (assuming 20bp gRNA length) to guide how much padding to add to your targets:
|Editing window size (gRNA length)||Expected INDEL length||Padding to add||Final target size|
|20 bp||<= 5 bp||10 bp||40 bp|
|20 bp||6 - 10 bp||15 bp||50 bp|
|20 bp||11 - 20 bp||25 bp||70 bp|
|20 bp||21 - 30 bp||35 bp||90 bp|
What can be done if one of my on-target sites cannot be covered by an amplicon using Tapestri Designer?
You can contact our sales or support teams about our White Glove service. Our White Glove team may be able to perform a ‘rescue’ of the target. Additionally, we can try adding this rescued amplicon into the panel at a higher concentration in an attempt to improve performance. However, rescued amplicons are known to have lower performance, therefore we additionally recommend sequencing at a very high depth to retrieve as much data as possible.
How can I determine my off-target sites?
There are several predictive tools for potential off-targets. One free academic tool available is CHOPCHOP.
What are the sequencing depth recommendations for the Genome Editing pipeline?
Our sequencing recommendations are the same as for our standard DNA + Protein pipeline, which you can read more about in this article.