空间转录组测序学习-样本准备和建库

内容来自视频，网址附后

主要内容如下

Goals Agenda
why
why-2
product-definitions
Visium Spatial Tissue Optimization Workflow Visium Spatial Tissue Optimization Workflow-2 Visium Spatial tissue Optimization Kit
How to choose the best permeabilization time Consider the following factors with tissue optimization
examle of fluorescent cDNA footprint
Optimal permeabilization time determined from tissue optimization proportionally correlates with maximum sensitivity
Tissue Optimization Points summary
Purpose for Each Step
Unbiased Gene Expression at high spatial resolution
On slide - off slide

Tissue Prepartaion and Compatibility

Tissue Preparation-Freezing and Embedding
Cryosectioning and Tissue Placement
Tissue Preparation - Proper Tissue Placement and Handling
Cryosectioning Thickness Practice Sectioning Visium Tissue Compatibility
https://support.10xgenomics.com/spatial-gene-expression/tissue-optimization/doc/specifications-visium-spatial-gene-expression-optimized-tissues Current tissues conditios Preparing for Imaging of Visium Slides imaging guide line

Sequencing

空间转录组学-冷冻样品 单细胞测序-过程 Gene Expression Library
Visium Gene Expression Library

Minimum Sequencing Depth: We recommend a minimum of 50k read pairs per spot covered with tissue. To calculate this, first estimate the % of capture area covered by the tissue section based upon the H&E brightfield image. Then use the following formula to calculate the recommended sequencing depth:
• Total sequencing depth = (Coverage Area x total spots on the Capture Area) x 50,000 read pairs/spot
Example calculation for 60% tissue coverage: (0.60 x 5,000 total spots) x 50,000 read pairs/spot = 150 million total read pairs for that sample
Additional guidelines for estimating the number of spots covered: Determining number of spots covered with tissue may be performed visually or by using the Visium Manual Alignment Wizard in Loupe Browser for a more accurate measurement.

https://support.10xgenomics.com/spatial-gene-expression/sequencing/doc/specifications-sequencing-requirements-for-visium-spatial-gene-expression
https://support.10xgenomics.com/spatial-gene-expression/software/pipelines/latest/what-is-space-ranger