Question: What library types can be pooled prior to hybridization in the Targeted Gene Expression assay?
Answer: The Targeted Gene Expression assay generates targeted libraries from Whole Transcriptome Analysis (WTA) input libraries. Supported input library types include:
- 3’ Single Cell Gene Expression libraries (v3, v3.1 and dual index v3.1)
- 5’ Single Cell Gene Expression libraries (v1, v1.1 and v2)
- Visium Spatial Gene Expression libraries (prepared using Rev D* of CG000239)
For the Targeted Gene Expression assay, up to eight WTA input libraries can be pooled prior to hybridization.
We recommend pooling input libraries of the same library type (Single Cell 3' / Single Cell 5' / Spatial) with the same indexing strategy (single index / dual index), prepared from similar cell/tissue types and similar cell/spot numbers:
- While pooling libraries from different cell types and pooling Single Cell 3' and 5' Gene Expression libraries is compatible with the Targeted Gene Expression assay, it can lead to sequencing read imbalance.
- Pooling single index and dual index libraries can introduce additional complexity due to varying read configurations required for sequencing.
- We do not support or recommend pooling Single Cell 3' or 5' Gene Expression libraries with Visium Spatial Gene Expression libraries
Do not pool libraries with the same sample index, as pooled libraries will be sequenced together. For adequate sample demultiplexing post-sequencing, each library must contain a unique sample index.
Products: Targeted Gene Expression