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  1. IGB
  2. IGBF-3863

Investigate Seurat marker gene alignments in IGB

        Details

        • Type: Task
        • Status: Closed (View Workflow)
        • Priority: Major
        • Resolution: Done
        • Affects Version/s: None
        • Fix Version/s: None
        • Labels:
          None

          Description

          In IGBF-3838, the PBMC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) did not line up with the reference gene models. The reads are misaligned from the boundaries of the exons in the gene model. The UMI count matrix data for the PBMC dataset was used in the Seurat Guided Clustering Tutorial . To investigate further, view the the reads of the the marker genes mentioned in the tutorial in IGB and check how many are aligned with the gene model.

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              Task - A task that needs to be done. IGBF-3864 Find more single cell RNA seq datasets in10XGenomics

              • Major - Major loss of function.
              • Closed

              Task - A task that needs to be done. IGBF-3838 Obtain and view sequence read alignments for PBMC dataset from Seurat vignette

              • Major - Major loss of function.
              • Closed

                Activity

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                karthik Karthik Raveendran created issue -
                karthik Karthik Raveendran made changes -
                Field Original Value New Value
                Epic Link IGBF-3765 [ 22984 ]
                karthik Karthik Raveendran made changes -
                Status To-Do [ 10305 ] In Progress [ 3 ]
                karthik Karthik Raveendran made changes -
                Link This issue is blocked by IGBF-3864 [ IGBF-3864 ]
                karthik Karthik Raveendran made changes -
                Link This issue relates to IGBF-3838 [ IGBF-3838 ]
                karthik Karthik Raveendran made changes -
                Link This issue relates to IGBF-3864 [ IGBF-3864 ]
                karthik Karthik Raveendran made changes -
                Link This issue is blocked by IGBF-3864 [ IGBF-3864 ]
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Some of the genes mentioned in the ticket does not seem align with the reference genome. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are edge matched.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file.                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Some of the genes mentioned in the ticket does not seem align with the reference genome. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are edge matched.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
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                karthik Karthik Raveendran added a comment - - edited

                I was counting the reads that align within the exon boundaries of the gene model and recording it in a spreadsheet. After talking to Dr. Nowlan Freese, I realized I have been counting only reads with no gaps and did not count the ones with some gaps. The counts needs to be updated with counts of reads that may have some gaps but are still within the exon boundaries.

                We also discussed about the CellRanger pipeline and understanding how the UMI counts are counted for each feature (gene) for each cell or rather what reads are counted and what is discarded.

                Show
                karthik Karthik Raveendran added a comment - - edited I was counting the reads that align within the exon boundaries of the gene model and recording it in a spreadsheet. After talking to Dr. Nowlan Freese , I realized I have been counting only reads with no gaps and did not count the ones with some gaps. The counts needs to be updated with counts of reads that may have some gaps but are still within the exon boundaries. We also discussed about the CellRanger pipeline and understanding how the UMI counts are counted for each feature (gene) for each cell or rather what reads are counted and what is discarded.
                Hide
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                karthik Karthik Raveendran added a comment -

                10XGenomics has their own browser called Loupe browser. The browser can not be used to visualize bam files, as far as I can tell, but just clustering visualizations. Looking into this browser might be useful. However, the input files to the browser is a cloupe file and I could not find one for the dataset we are looking at now. This may be set aside for another time.

                Show
                karthik Karthik Raveendran added a comment - 10XGenomics has their own browser called Loupe browser. The browser can not be used to visualize bam files, as far as I can tell, but just clustering visualizations. Looking into this browser might be useful. However, the input files to the browser is a cloupe file and I could not find one for the dataset we are looking at now. This may be set aside for another time.
                Hide
                Permalink
                karthik Karthik Raveendran added a comment - - edited

                The prospect of adding a Samtags filter to IGB was also discussed, or at least a cell barcode filter will help in verifying the UMI count matrix in IGB.

                Show
                karthik Karthik Raveendran added a comment - - edited The prospect of adding a Samtags filter to IGB was also discussed, or at least a cell barcode filter will help in verifying the UMI count matrix in IGB.
                nfreese Nowlan Freese made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Some of the genes mentioned in the ticket does not seem align with the reference genome. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are edge matched.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Some of the genes mentioned in the ticket do not seem to align with the reference genome annotations (i.e. aligned RNA-Seq reads are falling within intronic regions, or are split across several genes). This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if the aligned reads are contained within the borders of the exons of the gene model.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Some of the genes mentioned in the ticket do not seem to align with the reference genome annotations (i.e. aligned RNA-Seq reads are falling within intronic regions, or are split across several genes). This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if the aligned reads are contained within the borders of the exons of the gene model.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Some of the genes mentioned in the ticket does not seem align with the reference genome. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Some of the genes mentioned in the ticket does not seem align with the reference genome. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. The single cell RNA sequences for the genes mentioned in the ticket does not seem to align with the the gene models of the reference genome. The reads are wildly misaligned with the edges of the each individual exons. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. The single cell RNA sequences for the genes mentioned in the ticket does not seem to align with the the gene models of the reference genome. The reads are wildly misaligned with the edges of the each individual exons. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. The single cell RNA sequences for the genes mentioned in the ticket does not seem to align with the the gene models of the reference genome. The reads are wildly misaligned with the edges of the each individual exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. The single cell RNA sequences for the genes mentioned in the ticket does not seem to align with the the gene models of the reference genome. The reads are wildly misaligned with the edges of the each individual exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. The single cell RNA sequences for the genes (mentioned in the ticket) does not seem to align with the the gene models of the reference genome. The reads are wildly misaligned with the edges of the each individual exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. The single cell RNA sequences for the genes (mentioned in the ticket) does not seem to align with the the gene models of the reference genome. The reads are wildly misaligned with the edges of the each individual exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. The single cell RNA sequences for the genes (mentioned in the ticket) does not seem to align with the reference gene models. The reads are wildly misaligned with the edges of the each individual exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. The single cell RNA sequences for the genes (mentioned in the ticket) does not seem to align with the reference gene models. The reads are wildly misaligned with the edges of the each individual exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. The single cell RNA sequences for the genes (mentioned in the ticket) does not seem to line up with the reference gene models. The reads are wildly misaligned with the edges of the exons in the gene model. If this is true for entire the bam file, then what are the counts of the UMI count matrix really about. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. The single cell RNA sequences for the genes (mentioned in the ticket) does not seem to line up with the reference gene models. The reads are wildly misaligned with the edges of the exons in the gene model. If this is true for entire the bam file, then what are the counts of the UMI count matrix really about. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) does not seem to line up with the reference gene models. The reads are wildly misaligned with the edges of the exons in the gene model. If this is true for entire the bam file, then what are the counts of the UMI count matrix really about. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) does not seem to line up with the reference gene models. The reads are wildly misaligned with the edges of the exons in the gene model. If this is true for entire the bam file, then what are the counts of the UMI count matrix really about. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they are aligned.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) does not seem to line up with the reference gene models. The reads are wildly misaligned with the edges of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) does not seem to line up with the reference gene models. The reads are wildly misaligned with the edges of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) does not line up with the reference gene models. The reads are wildly misaligned with the edges of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) does not line up with the reference gene models. The reads are wildly misaligned with the edges of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) does not line up with the reference gene models. The reads are wildly misaligned from the boundaries of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) does not line up with the reference gene models. The reads are wildly misaligned from the boundaries of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) do not line up with the reference gene models. The reads are wildly misaligned from the boundaries of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) do not line up with the reference gene models. The reads are wildly misaligned from the boundaries of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) did not line up with the reference gene models. The reads are misaligned from the boundaries of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) did not line up with the reference gene models. The reads are misaligned from the boundaries of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the marker genes (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) did not line up with the reference gene models. The reads are misaligned from the boundaries of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the sequences of the (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).
                karthik Karthik Raveendran made changes -
                Description In IGBF-3838, the PMBC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) did not line up with the reference gene models. The reads are misaligned from the boundaries of the exons in the gene model. This data was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, look at the sequences of the (mentioned in the tutorial) in IGB and check if they line up.

                Note: the Seurat tutorial does not use the bam file for their analysis. They use a UMI count matrix file (output of [CellRanger | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger] ).                 		In IGBF-3838, the PBMC dataset of 2700 single cells was obtained and viewed in IGB. Almost all of the single cell RNA sequences for the genes (mentioned in the ticket) did not line up with the reference gene models. The reads are misaligned from the boundaries of the exons in the gene model. The UMI count matrix data for the PBMC dataset was used in the [Seurat Guided Clustering Tutorial | https://satijalab.org/seurat/articles/pbmc3k_tutorial.html]. To investigate further, view the the reads of the the marker genes mentioned in the tutorial in IGB and check how many are aligned with the gene model.
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                karthik Karthik Raveendran added a comment -

                Images of the PBMC reads and respective depth graphs are added here: https://drive.google.com/drive/u/0/folders/1Bh2zG9UYoeFEGmYohk1c5-gFCRUE-zC4

                Note: Depth graphs images have "DG" at the end of their filenames

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                karthik Karthik Raveendran added a comment - Images of the PBMC reads and respective depth graphs are added here: https://drive.google.com/drive/u/0/folders/1Bh2zG9UYoeFEGmYohk1c5-gFCRUE-zC4 Note: Depth graphs images have "DG" at the end of their filenames
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                karthik Karthik Raveendran added a comment -

                The coverage of the ranges from 2.5k to 70k reads and 3' bias was observed on all marker genes. This dataset uses cellranger v1.1.0 and chemistry v1.0. Datasets created with newer cellranger and chemistry versions need to be found and observed (IGBF-3864). Bulk RNA seq should also be visualized for comparison. Moving this ticket to close.

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                karthik Karthik Raveendran added a comment - The coverage of the ranges from 2.5k to 70k reads and 3' bias was observed on all marker genes. This dataset uses cellranger v1.1.0 and chemistry v1.0. Datasets created with newer cellranger and chemistry versions need to be found and observed ( IGBF-3864 ). Bulk RNA seq should also be visualized for comparison. Moving this ticket to close.
                karthik Karthik Raveendran made changes -
                Status In Progress [ 3 ] Needs 1st Level Review [ 10005 ]
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                Status Needs 1st Level Review [ 10005 ] First Level Review in Progress [ 10301 ]
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                Status First Level Review in Progress [ 10301 ] Ready for Pull Request [ 10304 ]
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                Status Ready for Pull Request [ 10304 ] Pull Request Submitted [ 10101 ]
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                Status Pull Request Submitted [ 10101 ] Reviewing Pull Request [ 10303 ]
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                Status Reviewing Pull Request [ 10303 ] Pull Request Submitted [ 10101 ]
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                Status Pull Request Submitted [ 10101 ] Reviewing Pull Request [ 10303 ]
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                Status Reviewing Pull Request [ 10303 ] Merged Needs Testing [ 10002 ]
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                Status Merged Needs Testing [ 10002 ] Post-merge Testing In Progress [ 10003 ]
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                Resolution Done [ 10000 ]
                Status Post-merge Testing In Progress [ 10003 ] Closed [ 6 ]

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                  • Assignee:
                    karthik Karthik Raveendran
                    Reporter:
                    karthik Karthik Raveendran
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