FAQs

Kits

What is included in a kit?


Everything you need to go from a single-cell or single-nuclei suspension to a ready-to-sequence library. We also provide a computational pipeline that processes your raw sequencing data to a cell-gene matrix that integrates into downstream open source analysis tools along with an experimental summary report.




What prior equipment do I need to run a kit?


The primary equipment required are multichannel pipettes, a thermocycler, and a swinging-bucket centrifuge.




What is a sublibrary?


After the third round barcoding and before lysis, cells will be pooled and subsequently split into between one and eight distinct populations we call sublibraries. Each sublibrary will contain a different 4th barcode (the Illumina index) and can be sequenced separately. You will count cells before splitting them into your sublibraries, and thus you can choose the number of cells in each. In practice it can be useful to have at least one sublibrary with few cells, which can provide an estimate of gene detection per cell at deeper sequencing while using few sequencing reads.




Do I need to purchase a custom instrument to run a Split Bio kit?


No, our kits are standalone and do not require any custom instrument.




Can I do single nucleus transcriptome sequencing?


Yes, our protocol is compatible with nuclei. We offer two separate fixation kits for work with cells or nuclei.




Is FACS sorting required?


Our protocols do not require FACS sorting.




What kind of sequencing kit is required?


Split Bio kits use paired-end 2x75 cycle sequencing kits.




Does the Single Cell Whole Transcriptome kit use standard Illumina primers?


Yes, the resulting library can be loaded into Illumina sequencers using standard protocols (no custom primers).





Sequencing

Does the Single Cell Whole Transcriptome kit use standard Illumina primers?


Yes, the resulting library can be loaded into Illumina sequencers using standard protocols (no custom primers).




Can I sequence a subset of cells from a single experiment?


Yes, you can aliquot anywhere from 100 to 12,500 barcoded cells into different sublibraries. It is possible to sequence a single sublibrary independently. You can deeply sequence a small cell sublibrary (i.e. 100 cells) and understand gene detection before proceeding to large cell sublibraries (i.e. 12,500 cells).




What instrument should I use for sequencing?


Libraries are compatible with all Illumina sequencing instruments.





Method

Can I do single nucleus transcriptome sequencing?


Yes, our protocol is compatible with nuclei. We offer two separate fixation kits for work with cells or nuclei.




Is FACS sorting required?


Our protocols do not require FACS sorting.




Can I preserve my samples before doing my single-cell experiment?


Yes, upon extracting your sample, you will perform fixation (30 min to 1 hour). After this you can store your samples for up to 6 months before proceeding to our barcoding and library preparation protocol. This makes it possible to pool samples extracted from different days into a single run.




What is a sublibrary?


After the third round barcoding and before lysis, cells will be pooled and subsequently split into between one and eight distinct populations we call sublibraries. Each sublibrary will contain a different 4th barcode (the Illumina index) and can be sequenced separately. You will count cells before splitting them into your sublibraries, and thus you can choose the number of cells in each. In practice it can be useful to have at least one sublibrary with few cells, which can provide an estimate of gene detection per cell at deeper sequencing while using few sequencing reads.




Can I sequence a subset of cells from a single experiment?


Yes, you can aliquot anywhere from 100 to 12,500 barcoded cells into different sublibraries. It is possible to sequence a single sublibrary independently. You can deeply sequence a small cell sublibrary (i.e. 100 cells) and understand gene detection before proceeding to large cell sublibraries (i.e. 12,500 cells).





Data Analysis

How do I process my data?


Split Bio provides a pipeline that can be run locally on your server. The pipeline takes fastq files and delivers processed data in the form of a cell-gene count matrix, which serves as the input for various open sources tools such as scanpy and seurat. The pipeline also delivers an experimental summary report with run statistics.




Can I sequence a subset of cells from a single experiment?


Yes, you can aliquot anywhere from 100 to 12,500 barcoded cells into different sublibraries. It is possible to sequence a single sublibrary independently. You can deeply sequence a small cell sublibrary (i.e. 100 cells) and understand gene detection before proceeding to large cell sublibraries (i.e. 12,500 cells).




How do I know how my run went?


Split Bio’s pipeline will deliver an experimental summary report with run statistics. These include the number of detected cells, gene and UMI detection, sequencing quality, and a few other metrics relevant to your experiment.





All

Do I need to purchase a custom instrument to run a Split Bio kit?


No, our kits are standalone and do not require any custom instrument.




What prior equipment do I need to run a kit?


The primary equipment required are multichannel pipettes, a thermocycler, and a swinging-bucket centrifuge.




What is included in a kit?


Everything you need to go from a single-cell or single-nuclei suspension to a ready-to-sequence library. We also provide a computational pipeline that processes your raw sequencing data to a cell-gene matrix that integrates into downstream open source analysis tools along with an experimental summary report.




Can I do single nucleus transcriptome sequencing?


Yes, our protocol is compatible with nuclei. We offer two separate fixation kits for work with cells or nuclei.




Is FACS sorting required?


Our protocols do not require FACS sorting.




What is a sublibrary?


After the third round barcoding and before lysis, cells will be pooled and subsequently split into between one and eight distinct populations we call sublibraries. Each sublibrary will contain a different 4th barcode (the Illumina index) and can be sequenced separately. You will count cells before splitting them into your sublibraries, and thus you can choose the number of cells in each. In practice it can be useful to have at least one sublibrary with few cells, which can provide an estimate of gene detection per cell at deeper sequencing while using few sequencing reads.




Does the Single Cell Whole Transcriptome kit use standard Illumina primers?


Yes, the resulting library can be loaded into Illumina sequencers using standard protocols (no custom primers).




Can I preserve my samples before doing my single cell experiment?


Yes, upon extracting your sample, you will perform fixation (30 min to 1 hour). After this you can store your samples for up to 6 months before proceeding to our barcoding and library preparation protocol. This makes it possible to pool samples extracted from different days into a single run.




Can I sequence a subset of cells from a single experiment?


Yes, you can aliquot anywhere from 100 to 12,500 barcoded cells into different sublibraries. It is possible to sequence a single sublibrary independently. You can deeply sequence a small cell sublibrary (i.e. 100 cells) and understand gene detection before proceeding to large cell sublibraries (i.e. 12,500 cells).




What instrument should I use for sequencing?


Libraries are compatible with all Illumina Sequencing instruments.




Does the Single Cell Whole Transcriptome Kit use standard Illumina primers?


Yes, the resulting library can be loaded into Illumina sequencers using standard protocols (no custom primers).




How do I process my data?


Split Bio provides a pipeline that can be run locally on your server. The pipeline takes fastq files and delivers processed data in the form of a cell-gene count matrix, which serves as the input for various open sources tools such as scanpy and seurat. The pipeline also delivers an experimental summary report with run statistics.




How do I know how my run went?


Split Bio’s pipeline will deliver an experimental summary report with run statistics. These include the number of detected cells, gene and UMI detection, sequencing quality, and a few other metrics relevant to your experiment.





Split Bio offers a new strategy for single-cell RNA sequencing. Our pioneering technology, known as SPLiT-seq (Split-pool ligation-based transcriptome sequencing), uses combinatorial cDNA barcoding within cells themselves, and thus does not require complex cell partition instruments. Our Single Cell Whole Transcriptome kit has improved on the SPLiT-seq method and enables profiling of up to one hundred thousand unique cells in a single experiment. 

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