Spatiotemporal transcriptomic atlas of mouse organogenesis using DNA nanoball-patterned arrays

May combine two claims. The quote only partly states this claim.

The epithelial cell cluster (Epcam+, Krt8+) could be grouped into specialized epithelial cell subtypes including hair follicle cells, alveolar type II cells, club cells, ciliated cells, zona glomerulosa epithelial cells, glandular epithelium, stratified squamous epithelium, renal tubular cells, and thymic epithelial cells.

“For example, the epithelial cell cluster ( Epcam + , Krt8 + ) could be grouped into specialized epithelial cell subtypes including hair follicle cells ( Krt5 + , Krt17 + ), alveolar type II cells ( Foxp2 + , Nkx2-1 + ), club cells ( Scgb3a2 + , S100a6 + ), and ciliated cells ( Dynlrb2 + , Foxj1 + ) from the lung”

May combine two claims.

There was substantial colocalization of nucleic acid dye and aggregated transcripts detected by Stereo-seq, illustrating that cells can be segmented in an image-dependent manner.

“There was substantial colocalization of the dye and the aggregated transcripts detected by Stereo-seq (Figures 2A and 2B), illustrating that cells can be segmented in an image-dependent manner.”

May combine two claims.

Image-based cell segmentation yielded 50,140 segmented cells with an average of 1,910 UMI and 792 genes per cell.

“After filtering out the low-capture cells, we obtained 50,140 segmented cells with an average of 1,910 UMI and 792 genes per cell (Figure S1F).”

May combine two claims.

For E9.5-E15.5 stages, four to six sections were included from different embryos; for E16.5, 18 sections were profiled, with a series of 13 sections from one single embryo and five from another to assess reproducibility, allowing coverage of all major tissues and organs.

“For E9.5-E15.5 stages, four to six sections were included from different embryos. For E16.5, 18 sections were profiled, with a series of 13 sections from one single embryo and five from another to assess reproducibility, allowing coverage of all major tissues and organs (Table S1).”

May combine two claims.

Among the E16.5 regulons, ARNT2 was enriched in the brain, FOXC2 in the meninges, GATA4 in the heart, SOX9 in the bone, and MEF2C in the muscle.

“Among the E16.5 regulons, ARNT2 was enriched in the brain, FOXC2 in the meninges, GATA4 in the heart, SOX9 in the bone, and MEF2C in the muscle (Figure 3E).”

May combine two claims.

Stereo-seq has a larger number of spots per 100 μm2, with smaller spot size and center-to-center distance, than any other published method.

“Importantly, Stereo-seq has a larger number of spots per 100 m m 2 , with smaller spot size and center-to-center distance, than any other published method (Figure 1B).”

May combine two claims.

At E9.5, 31 modules containing a total of 498 regulons were identified, and at E16.5, 35 modules containing a total of 469 regulons that correspond to different tissues.

“For example, at E9.5, we identified 31 modules containing a total of 498 regulons, and at E16.5, we identified 35 modules containing a total of 469 regulons that correspond to different tissues (Figures 3D, S4F, and S4G).”

May combine two claims.

In the E13.5 spinal cord, Stereo-seq identified the ventricular zone (Hopx+), marginal zone (Slc5a7+), basal plate (Vsnl1+), ventral (Fut9+) and lateral parts of the spinal alar plate (Hoxb8+), and superficial stratum of spinal basal plate (Pdyn+); in the E16.5 kidney, it localized artery (Myl9+), cortex (Lhx1+), medulla (Nrxn3+), capsule (Robo2+), and glomeruli (Poxdl+).

“For example, in the E13.5 spinal cord, we identified the ventricular zone ( Hopx + ), marginal zone ( Slc5a7 + ), basal plate ( Vsnl1 + ), ventral ( Fut9 + ) and lateral parts of the spinal alar plate ( Hoxb8 + ), and superficial stratum of spinal basal plate ( Pdyn + ); in the E16.5 kidney, we localized artery ( Myl9 + ), cortex ( Lhx1 + ), medulla ( Nrxn3 + ), capsule ( Robo2 + ), and glomeruli”

May combine two claims.

Stereo-seq combines single-cell resolution, high sensitivity, and a large field of view.

“Here, we report the development of a DNB-based genome-wide technology, Stereo-seq, that combines single-cell resolution, high sensitivity, and a large field of view”

May combine two claims.

The average number of captured genes per bin ranged from 1,770 at E15.5 to 3,900 at E10.5 and of UMI from 4,357 at E15.5 to 13,789 at E9.5.

“Theaveragenumberofcapturedgenes per bin ranged from 1,770 at E15.5 to 3,900 at E10.5 and of UMI from 4,357 at E15.5 to 13,789 at E9.5 (Figure S2A; Table S1).”

The quote only partly states this claim.

Spatial reclustering of bins from areas corresponding to selected tissues revealed detailed subregions, including in E13.5 spinal cord and E16.5 kidney.

“In addition, since Stereo-seq captures high density of signals, we performed spatial reclustering of bins from areas corresponding to selected tissues, which revealed detailed subregions.”

Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing).

Reclustering of specific cell populations could further identify cell subtypes or cell states.

“Reclustering of specific cell populations could further identify cell subtypes or cell states.”

May combine two claims.

After filtering out low-capture cells, the E16.5 embryo section yielded 281,377 segmented cells with an average of 1107 UMI and 529 genes per cell.

“After filtering out the low-capture cells, we obtained 281,377 segmented cells with an average of 1107 UMI and 529 genes per cell (Figures 4A and S5A).”

May combine two claims.

The 25 major cell types were either located in specific anatomic regions or represented common cell types spread across multiple sites.

“These cell types were either located in specific anatomic regions (e.g., cardiomyocytes, hepatocytes, and different types of neurons) or represented common cell types spread across multiple sites (e.g., chondrocytes, epithelial cells, myoblasts, and endothelial cells).”

Probably not a substantial claim: may not pass S3 (load-bearing).

The distribution of genes and UMI per bin varied between sections from different time points.

“Thedistribution of genes and UMI perbin varied between sections from different time points (Figure S2B).”

May combine two claims.

Stereo-seq chips of up to an effective area of 13.2 cm × 13.2 cm have been used for different tissue sizes, subdivided into 50, 100, and 200 mm2 chips for profiling sections from mouse olfactory bulb, mouse hemibrain, and whole mouse embryos.

“So far, we have used Stereo-seq chips of up to an effective area of 13.2 cm 3 13.2 cm for different tissue sizes, subdivided into 50, 100, and 200 mm 2 chips for profiling sections from the mouse olfactory bulb ( 10.5 mm 2 ), a mouse hemibrain ( 24.2 mm 2 ) and whole mouse embryos ( 7.1 to 76.1 mm 2 ), respectively (Figure S1A).”

May combine two claims.

The distribution of specific markers (Pcp4 and Slc17a7) captured by Stereo-seq exhibited the expected similarity to in situ hybridization data from Allen Brain Atlas and clearer patterns than HDST or Slide-seqV2-resolved expression data.

“The distribution of specific markers ( Pcp4 and Slc17a7 ) captured by Stereo-seq exhibited the expected similarity to in situ hybridization (ISH) data taken from Allen Brain Atlas (ABA) (Lein et al., 2007) and clearer patterns than HDST (Vickovic et al., 2019) or Slide-seqV2-resolved (Stickels et al., 2021) expression data (Figure S1E).”

• cites Lein et al., 2007 sourceNot checked yet

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• cites Vickovic et al., 2019 sourceNot checked yet

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• cites Stickels et al., 2021Not checked yet

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May combine two claims.

Gene programs identified by Hotspot showed well-defined spatial patterns that largely match anatomic regions and contained sets of genes that could be validated by ISH.

“The resulting gene programs showed well-defined spatial patterns that, to a large extent, match anatomic regions and contained sets of genes that could be validated by ISH (e.g., Kitl colocalizing with Shh in the E9.5 notochord and Pantr1 with Sox2 in the E11.5 brain) (Figures S4A-S4E).”

• cites DeTomaso and Yosef, 2021Not checked yet

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• AssumptionRests on an assumption: Amalgamation of different signals in complex tissues can complicate interpretations.

May combine two claims.

Standard DNB chips have spots with approximately 220 nm diameter and a center-to-center distance of 500 or 715 nm, providing up to 400 spots for tissue RNA capture per 100 μm2.

“Standard DNB chips have spots with approximately 220 nm diameter and a center-to-center distance of 500 or 715 nm (Figure 1A, step 1), providing up to 400 spots for tissue RNA capture per 100 m m 2 .”

May combine two claims.

High Wnt5a expression was observed in mesenchymal cells and fibroblasts from the maxilla, whereas in the limb it was located mainly in mesenchymal cells.

“We observed high Wnt5a expression in mesenchymal cells and fibroblasts from the maxilla, whereas in the limb, it was located mainly in mesenchymal cells (Figures 6C and 6D).”

May combine two claims.

HES6 was highly active in neuroblasts, and its target genes included Ascl1 and Hes5.

“HES6 was highly active in neuroblasts, and its target genes included Ascl1 and Hes5 (Figures 5L and S7G).”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Unknown potential regulators Ttc3 and Celf4 of interneuron migration were observed and could be interesting for further exploration.

“We also observed unknown potential regulators ( Ttc3 and Celf4 ) of interneuron migration that could be interesting for further exploration (Figure 4F).”

May combine two claims.

Highly specific expressions of extrinsic cues (ligands) were identified in glioblasts (e.g., Rspo1/2/3, Wnt1, Wnt9a) and neuroblasts (Fgf15, Wnt7b, Tnc), providing potential mechanistic insight for further specification of these progenitors.

“In addition, we identified highly specific expressions of extrinsic cues (ligands) in glioblasts (e.g., Rspo1/2/3 , Wnt1 , and Wnt9a ) and neuroblasts ( Fgf15, Wnt7b, and Tnc ) (Figures 5G-5I and MOSTA website), providing potential mechanistic insight for the further specification of these progenitors.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Stereo-seq is an affordable technology.

“Moreover, Stereo-seq is an affordable technology.”

May combine two claims.

At E14.5, radial glia cells disappeared from the rostral part and remained in the more caudal domains of the ventricular zone.

“At E14.5, the radial glia cells disappeared from the rostral part and remained in the more caudal domains of the ventricular zone, revealing the spatiotemporal development of this brain region.”

May combine two claims.

Wnt5a localized in the craniofacial region, limb, genital ridge, and brain (floor plate and neocortex) from multiple developmental stages, consistent with the disease phenotype.

“We observed that Wnt5a localized in the craniofacial region, limb, genital ridge, and brain (floor plate and neocortex) from multiple developmental stages (Figure 6B), consistent with the disease phenotype (Menezes et al., 2010).”

• cites Menezes et al., 2010Not checked yet

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May combine two claims.

Developmental trajectories revealed two branches arising from radial glia cells, one to neuroblasts and the other to glioblasts.

“The resulting developmental trajectories revealed two branches arising from radial glia cells, one to neuroblasts andthe other to glioblasts (Figure 5D).”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

The high resolution of Stereo-seq allows efficient image-based cell segmentation, but in some cases where multiple cell types are close to each other, particularly smaller cell types like immune cells, cell segmentation may be imperfect.

“The high resolution of Stereo-seq allows efficient image-based cell segmentation, but in some cases, where multiple cell types are close to each other, particularly smaller cell types like immune cells, cell segmentation may be imperfect.”

May combine two claims.

HES1 was highly active in glioblasts, and its target genes included known glia-lineage genes such as Aldh1l1 and Nfib.

“For example, HES1 was highly active in glioblasts, and its target genes included known genes specific to the glia lineage such as Aldh1l1 and Nfib .”

May combine two claims.

Radial glia cells (Mki67+, Dbx1+), the source of both neuroblasts and glioblasts, were the most abundant progenitors at E12.5, with widespread distribution.

“Radial glia cells ( Mki67 + , Dbx1 + ), which are the source of both neuroblasts ( Npas3 + , Adgrb3 + ) and glioblasts ( Aldh1l1 + , Efna5 + ), were the most abundant progenitors at E12.5, with widespread distribution.”

May combine two claims.

The MSX1 regulon was more enriched in mesenchymal cells and fibroblasts in the maxilla, and mesenchymal cells in the limb.

“Similarly, the MSX1 regulon was more enriched in mesenchymal cells and fibroblasts in the maxilla, and mesenchymal cells in the limb (Figures 6C and 6D).”

May combine two claims.

Msx1 was expressed at different levels in multiple cell types of the maxilla including mesenchymal cells, but in the limb it was rather restricted to mesenchymal cells only.

“Msx1 was expressed at different levels in multiple cell types of the maxilla including mesenchymal cells, but in the limb, it was rather restricted to the latter cell type only.”

Classified as probably not a substantial claim. May combine two claims.

The current DNB capture chip costs 220 RMB/mm2 ($35/mm2) or 22,000 RMB/cm2 ($3,500/cm2).

“The current DNB capture chip costs 220 RMB/mm 2 ($35/ mm 2 ) or 22,000 RMB/cm 2 ($3,500/cm 2 ).”

May combine two claims.

Many disease-related genes were mainly enriched in one tissue or system, while others were more widely distributed.

“Similarly, although many genes were mainly enriched in one tissue (in some cases, they were confined to a specific region) or system, others were more widely distributed.”

May combine two claims.

The E16.5 Stereo-seq map identified expression of interneuron progenitor markers (Dlx1+, Dlx2+, Lhx6+, Nkx2-1+) within the medial ganglionic eminence and the migratory trajectory of interneurons into the cerebral cortex.

“Our E16.5 Stereo-seq map clearly identified the expression of interneuron progenitor markers ( Dlx1 + , Dlx2 + , Lhx6 + , Nkx2-1 + ) (Lim et al., 2018) within the medial ganglionic eminence (MGE) and the migratory trajectory of interneurons into the cerebral cortex (Figures 4D and S6D).”

• cites Lim et al., 2018 sourceNot checked yet

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• AssumptionRests on an assumption: Dorsal cortical ventricular zones give rise to glutamatergic neurons that populate the cerebral cortex and hippocampus, and ventral ventricular zones of the lateral, medial, and caudal ganglionic eminence give rise to GABAergic interneurons that populate the striatum and globus pallidus and migrate into the cerebral cortex.

May combine two claims.

Over 10% of Stereo-seq captured transcripts contained intronic reads, comparable with the proportion captured by scRNA-seq.

“In this regard, over 10% of the Stereo-seq captured transcripts contained intronic reads (Figure S6E), comparable with the proportion captured by scRNA-seq (La Manno et al., 2018; Qiu et al., 2022b).”

• cites La Manno et al., 2018 sourceNot checked yet

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• cites Qiu et al., 2022bNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

Stereo-seq has genome-wide coverage, but due to limitations in capture, relevant genes with low expression may be missed.

“Similarly, Stereo-seq has a genome-wide coverage, but due to limitations in capture, relevant genes with low expression may be missed.”

May combine two claims.

Some disease-related genes were expressed in most or all embryonic stages, whereas others had a restricted window.

“Some genes were expressed in most or all embryonic stages, whereas others had a restricted window.”

May combine two claims.

At E12.5, there was only occasional appearance of sparsely distributed neuroblasts, whereas glioblasts localized mostly in the central and caudal parts.

“At this stage, there was only occasional appearance of sparsely distributed neuroblasts, whereas glioblasts localized mostly in the central and caudal parts.”

May combine two claims.

Although the function of many transcription factors controlling these regulons has been previously reported, the study identified the spatial domains in which they act.

“Although the function of many of the transcription factors controlling these regulons has been previously reported, we have identified the spatial domains in which they act.”

May combine two claims.

Stereo-seq also captures intronic transcripts, making it possible to calculate spatial RNA velocity with single-cell resolution, which can facilitate the study of cell fate transitions.

“Importantly, Stereo-seq also captures intronic transcripts, making it possible to calculate spatial RNA velocity with singlecell resolution, which as in scRNA-seq studies (La Manno et al., 2018; Qiu et al., 2022b), can facilitate the study of cell fate transitions.”

• cites La Manno et al., 2018 sourceNot checked yet

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• cites Qiu et al., 2022bNot checked yet

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May combine two claims.

NKX2-1 was mainly enriched in brain (hippocampus and subpallium) and lung, and Lmod3 was enriched in skeletal muscle.

“As informative examples, NKX2-1 , which is mutated in brainlung-thyroid syndrome (Shetty et al., 2014), was mainly enriched in brain (hippocampus and subpallium) and lung and Lmod3 , which is associated with nemaline myopathy (Berkenstadt et al., 2018), was enriched in the skeletal muscle (Figure 6B).”

• cites Shetty et al., 2014Not checked yet

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• cites Berkenstadt et al., 2018Not checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

The high-density RNA capture with DNB chip requires a sequencing depth of around 150 gigabases/cm2, which costs 5,000 RMB ($800) based on current short-read sequencing platforms.

“The high-density RNA capture with DNB chip requires a sequencing depth of around 150 gigabases/cm 2 , which costs 5,000 RMB ( $800) based on current short-read sequencing platforms.”

May combine two claims.

At E16.5, radial glia cells further decreased and concentrated in the caudal part, with neuroblasts showing a more restricted distribution and glioblasts expanding in number in the original locations.

“At E16.5, radial glia cells further decreased and concentrated in the caudal part, with neuroblasts showing a more restricted distribution and glioblasts expanding in number in the original locations.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Regulons with significant spatial autocorrelation were grouped into different modules based on pairwise spatial correlations of an E9.5 section.

“(F) Heatmap showing the regulons with significant spatial autocorrelation grouped into different modules based on pairwise spatial correlations of an E9.5 section (E1S1).”

Probably not a substantial claim: may not pass S3 (load-bearing).

Stereo-seq chips of different sizes range from 50 mm2 to 174.24 cm2.

“(A) Stereo-seq chips of different sizes ranging from 50 mm 2 to 174.24 cm 2 .”

Probably not a substantial claim: may not pass S3 (load-bearing).

Genes with significant spatial autocorrelation were grouped into different gene modules based on pairwise spatial correlations of E9.5 and E16.5 sections.

“(A and B) Heatmaps showing the genes with significant spatial autocorrelation grouped into different gene modules based on pairwise spatial correlations of E9.5 (left, E1S1) and E16.5 (right, E1S1) sections.”

May combine two claims.

The study profiled a total of 53 sagittal sections corresponding to an overall area of 23.4 cm2, but more sections may still be necessary to provide adequate coverage of all regional tissue types.

“Importantly, we have profiled a total of 53 sagittal sections corresponding to an overall area of 23.4 cm 2 , but more sections may still be necessary to provide adequate coverage of all regional tissue types.”

May combine two claims.

Stereo-seq spatially characterizes the transcriptomic organization and individual cell-type composition of complex tissues in an unbiased manner with high sensitivity and large field of view.

“Hence, Stereo-seq spatially characterizes the transcriptomic organization and individual cell-type composition of complex tissues in an unbiased manner with high sensitivity and large field of view.”

Classified as probably not a substantial claim.

Recent cell atlases using high-throughput scRNA-seq and scATAC-seq have provided an increasingly detailed view of mouse developmental gene expression and gene regulatory dynamics.

“Recent cell atlases using high-throughput single-cell RNA-sequencing (scRNA-seq) and single-cell assay for transposase accessible chromatin sequencing (scATAC-seq) technologies have provided an increasingly detailed view of mouse developmental gene expression and gene regulatory dynamics”

• cites Cao et al., 2019 sourceNot checked yet

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• cites He et al., 2020Not checked yet

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• cites Pijuan-Sala et al., 2020Not checked yet

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May combine two claims.

Stereo-seq data can be used to both identify functional anatomic regions in large tissues through bin-based clustering, and cell types based on image-guided cell segmentation.

“Therefore, Stereo-seq data can be used to both identify functional anatomic regions in large tissues through bin-based clustering, and cell types based on image-guided cell segmentation.”

Probably not a substantial claim: may not pass S3 (load-bearing).

The colocalization of Pantr1 with Sox2 is consistent with the regulatory function of this long noncoding RNA in early neuronal differentiation.

“The colocalization of Pantr1 with Sox2 is consistent with the regulatory function of this long noncoding RNA in early neuronal differentiation (Goff et al., 2015).”

• cites Goff et al., 2015Not checked yet

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May combine two claims.

Data can be extracted from scRNA-seq datasets using SCENIC, but the lack of spatial information is likely to confound important associations.

“These data can be extracted from scRNA-seq datasets using algorithms such as single-cell regulatory network inference and clustering (SCENIC) (Aibar et al., 2017), but the lack of spatial information is likely to confound important associations.”

• cites Aibar et al., 2017 sourceNot checked yet

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• AssumptionRests on an assumption: The lack of spatial information in scRNA-seq data is likely to confound important associations in gene regulatory network analysis.

Probably not a substantial claim: may not pass S3 (load-bearing).

Kitl is a pleiotropic factor that acts in the ontogeny of different cell types.

“Kitl is a pleiotropic factor that acts in the ontogeny of different cell types (Broudy, 1997).”

• cites Broudy, 1997Not checked yet

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Classified as probably not a substantial claim. May combine two claims.

Early mouse embryos, specific late-stage-embryonic tissues such as the cerebral cortex, and whole midgestation embryos have been analyzed using spatially resolved transcriptomic approaches.

“Early mouse embryos (Lohoff et al., 2022; Peng et al., 2019), specific late-stage-embryonic tissues such as the cerebral cortex (Di Bella et al., 2021; Stickels et al., 2021), and whole midgestation embryos (Liu et al., 2020; Srivatsan et al., 2021; Yao et al., 2020) have been analyzed using these approaches.”

• cites Lohoff et al., 2022Not checked yet

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• cites Di Bella et al., 2021Not checked yet

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• cites Liu et al., 2020 sourceNot checked yet

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Probably not a substantial claim: may not pass S4 (based).

Stereo-seq defines the spatiotemporal window of developmental disease vulnerability.

“Stereo-seq defines the spatiotemporal window of developmental disease vulnerability”

May combine two claims.

The results demonstrate the utility of MOSTA for defining spatiotemporal windows of disease-related gene expression during development and for establishing potential vulnerabilities.

“These results demonstrate the utility of MOSTA for defining the spatiotemporal windows of disease-related gene expression during development and for establishing potential vulnerabilities.”

Probably not a substantial claim: may not pass S4 (based). May combine two claims.

Stereo-seq is a DNB-based spatially resolved transcriptomic technology with genome-wide coverage, cellular resolution, high sensitivity, and large field of view.

“Stereo-seq is a DNB-based spatially resolved transcriptomic technology with genome-wide coverage, cellular resolution, high sensitivity, and large field of view.”

Probably not a substantial claim: may not pass S4 (based).

These parameters are all critical for accurately profiling the transcriptomic heterogeneity of relatively large mid- and late-stage mammalian embryos.

“These parameters are all critical for accurately profiling the transcriptomic heterogeneity of relatively large mid- and late-stage mammalian embryos.”

Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing), S4 (based).

Due to its characteristics and applicability, Stereo-seq also has the potential to move into routine clinical practice as a diagnostic tool complementary to medical imaging and histopathology data.

“Due to its characteristics and applicability, Stereo-seq also has the potential to move into routine clinical practice as an extraordinary diagnostic tool complementary to medical imaging and histopathology data.”

May combine two claims.

MOSTA comprehensively characterizes cell-type and cell-state heterogeneity at the whole embryo level, which is relevant for reconstructing cell differentiation processes involving regional and subregional specificities.

“MOSTA comprehensively characterizes cell-type and cellstate heterogeneity at whole embryo level, which is relevant for reconstructing cell differentiation processes involving regional and subregional specificities.”

• AssumptionRests on an assumption: The cell capture bias and lack of contextual microenvironment limits the power of scRNA-seq for predicting the origin and mechanisms of genetic diseases.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Msx1 is a homeobox gene encoding a transcription factor with known crosstalk with WNT5A.

“We then investigated Msx1 , a homeobox gene encoding a transcription factor with known crosstalk with WNT5A (Lin et al., 2011).”

• cites Lin et al., 2011Not checked yet

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Classified as probably not a substantial claim.

scRNA-seq datasets are becoming widely used to predict the origin and mechanisms of genetic diseases including developmental disorders.

“Nowadays, scRNA-seq datasets are becoming widely used to predict the origin and mechanisms of genetic diseases including developmental disorders (Watanabe et al., 2019).”

• cites Watanabe et al., 2019Not checked yet

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Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing), S4 (based).

Besides the study of development, Stereo-seq and its future refinements have the ability to transform multiple other research fields.

“Besides the study of development, Stereo-seq and its future refinements have the ability to transform multiple other research fields.”

Probably not a substantial claim: may not pass S3 (load-bearing).

The dorsal midbrain contains complex multilayered structures like the cerebral cortex that form the superior and inferior colliculi.

“The dorsal midbrain contains complex multilayered structures like the cerebral cortex that form the superior and inferior colliculi (Arimura et al., 2019).”

• cites Arimura et al., 2019Not checked yet

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Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing). May combine two claims.

Mice and humans differ in multiple aspects of normal physiology and disease; hence, there are caveats, but the approach clearly has the potential to uncover disease mechanisms.

“Mice and humans differ in multiple aspects of normal physiology and disease; hence, there are caveats, but this approach clearly has the potential to uncover disease mechanisms.”

• AssumptionRests on an assumption: Mice and humans differ in multiple aspects of normal physiology and disease.

May combine two claims.

The findings suggest an asynchronous and spatially heterogeneous model of neurogenesis and gliogenesis, with glioblasts appearing early at specific anatomic locations where they remain until later differentiation.

“These findings suggest an asynchronous and spatially heterogeneous model of neurogenesis and gliogenesis, with glioblasts appearing early at specific anatomic locations where they remain until later differentiation.”

• AssumptionRests on an assumption: The development of the dorsal midbrain has been less well characterized in terms of cellular taxonomy and organization compared with other brain regions.

Probably not a substantial claim: may not pass S3 (load-bearing).

All relevant procedures involving animal experiments presented in this study are compliant with ethical regulations regarding animal research.

“All relevant procedures involving animal experiments presented in this study are compliant with ethical regulations regarding animal research and were conducted under the approval of the Animal Care and Use committee of the Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences (license number IACUC2021002).”

Probably not a substantial claim: may not pass S3 (load-bearing).

Reads with MID containing either N bases or more than 2 bases with quality score lower than 10 were filtered out.

“Reads with MID containing either N bases or more than 2 bases with quality score lower than 10 were filtered out.”

Classified as probably not a substantial claim.

UMI with the same CID and the same gene locus were collapsed, allowing 1 mismatch to correct for sequencing and PCR errors.

“UMI with the same CID and the same gene locus were collapsed, allowing 1 mismatch to correct for sequencing and PCR errors.”

Probably not a substantial claim: may not pass S3 (load-bearing).

The whole Stereo-seq data processing procedure was integrated into a publicly available pipeline SAW.

“The whole procedure was integrated into a publicly available pipeline SAW available at https://github.com/BGIResearch/SAW.”

Probably not a substantial claim: may not pass S3 (load-bearing).

All relevant procedures involving animal experiments presented in this study were conducted under the approval of the Animal Care and Use committee of the Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences (license number IACUC2021002).

“All relevant procedures involving animal experiments presented in this study are compliant with ethical regulations regarding animal research and were conducted under the approval of the Animal Care and Use committee of the Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences (license number IACUC2021002).”

Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing), S4 (based).

Although algorithms including SCC were developed for the analyses in this study, others may be needed for a broader range of applications in various tissues from different species.

“Finally, although we have developed algorithms including the SCC for the analyses in this study, others may be needed for a broader range of applications in various tissues from different species.”

Classified as probably not a substantial claim.

For Figure 1C, to ensure proper comparisons, Stereo-seq data were binned into bin 3 (3x3 DNB, 2 mm), bin 14 (14x14 DNB, 10 mm) or bin 140 (140x140 DNB, 100 mm).

“For Figure 1C, to ensure that proper comparisons were made, the data of Stereo-seq were binned into bin 3 (3 3 3 DNB, 2 m m), bin 14 (14 3 14 DNB, 10 m m) or bin 140 (140 3 140 DNB, 100 m m).”

Classified as probably not a substantial claim.

CID sequences on the first reads were mapped to the designed coordinates of the in situ captured chip, allowing 1 base mismatch to correct for sequencing and PCR errors.

“CID sequences on the first reads were first mapped to the designed coordinates of the in situ captured chip achieved from the first round of sequencing, allowing 1 base mismatch to correct for sequencing andPCRerrors.”

Classified as probably not a substantial claim.

To validate cell type identity in the adult mouse hemibrain section, annotated scRNA-seq profiles of the adult mouse brain were used for comparison.

“To validate the cell type identity characterized in our adult mouse hemibrain section, annotated scRNA-seq profiles of the adult mouse brain downloaded from Linnarsson Lab website (http://mousebrain.org/adolescent/) (Zeisel et al., 2018) were used for comparison.”

• cites Zeisel et al., 2018 sourceNot checked yet

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