MAPseq/BARseq

This service provides 200-300 um cryostat sections for manual dissection purpose from fresh frozen tissue. Extra care is taken during cryostat sectioning to avoid cross sample contamination. The sections are stored at -80oC before dissection.

We are offering a free QC check on RNA quality and barcode amount for up to 10 mouse brain samples per lab. This service is available to labs from non-profit institutes between July 2024 and July 2027.

QC service includes:

• RNA extraction
• RIN check on a bioanalyzer
• RT-qPCR to examine barcode quantity

This free service is limited to 10 samples per lab.

Sequencing the MAPseq library by PE36 NextSeq

*Price per library

RNA extraction, QC for RNA quality and injection quality (bioanalyzer and q-PCR), Reverse Transcription and PCR to make the library for next generation sequencing.

RNA extraction, QC for RNA quality and injection quality (bioanalyzer and q-PCR), Reverse Transcription and PCR to make the library for next generation sequencing.

RNA extraction, QC for RNA and injection quality (bioanalyzer and q-PCR), Reverse Transcription and PCR to make the library for next generation sequencing.

RNA extraction, QC for RNA and injection quality (bioanalyzer and q-PCR), Reverse Transcription and PCR to make the library for next generation sequencing

RNA extraction, QC for RNA and injection quality (bioanalyzer and q-PCR), Reverse Transcription and PCR to make the library for next generation sequencing.

This package offers a comprehensive pilot study using both BARseq and MAPseq, designed to

help researchers generate preliminary data for grant applications or feasibility testing.

Included Services

1.Barcoded Sindbis Virus Library

o High-titer Sindbis virus: 10^10 ifu/mL

o Contains 40 million unique barcodes

o Supplied volume is sufficient to infect a 1 mm. brain area in at least 6 animals

o Developed and validated by the Zador lab for use in MAPseq and BARseq

2.Tissue Preparation

o Cryostat sectioning:

– 2 x 60 μm fixed sections for BARseq

– Up to 10 x 200 μm sections for MAPseq

o Manual dissection of up to 6 brain regions for MAPseq

o Dissection of 1 brain area, 2 sections for BARseq

3. BARseq Workflow

o One BARseq run on the selected region

o In situ sequencing:

– 7 cycles for gene sequencing

– 15 cycles for barcode sequencing

o Library preparation, rolony generation, and detection of barcode and 104 genes

4. MAPseq Workflow

o Library preparation for up to 6 dissected brain regions, including QC

o Pooled with other user sample for one high-throughput sequencing run

o Preliminary MAPseq data processing

Turnaround & Data Handling

• BARseq: The core provides the processing codes to users.

• MAPseq: Sequencing is pooled with other user samples; turnaround time depends on

demand.

Additional Notes

• Mouse brain injection and collection: Users are responsible for virus injection and

brain collection following the protocol provided by the core.

• Custom Probes: The package supports detection of 104 genes (Chen et al., Nature,

2024). Users may include additional probes at their own expense.

• Protocol Compliance: Deviations from the provided sample prep protocol may result in

extra charges.

Reference

Chen, X., Fischer, S., Rue, M.C.P. et al. Whole-cortex in situ sequencing reveals input-dependent area identity. Nature (2024). https://doi.org/10.1038/s41586-024-07221-6

• 5B=1.25B paired-end reads

We offer a preliminary Next Generation Sequencing (NGS) data processing service that includes the analysis of raw data and the generation of a Barcode Matrix, which provides information on the abundance of each barcoded neuron in each tissue sample.

Our pricing is based on a per sequencing run basis.

Starting from June 1st, 2023, our data processing service will undergo a significant update, resulting in a remarkable drop in the cost of data processing. Our new and improved pipeline is designed to be more efficient and faster, allowing us to deliver high-quality results at a much more competitive price point. This update represents our commitment to providing exceptional value to our users.

Run bioanalyzer to check RNA quality and RT-qPCR to examine barcode amount. Price listed is per 10 samples. RNA extraction is not included in the price.

Extract RNA from mice brain samples with Trizol reagent.

Provide Sindbis virus library containing millions of barcodes with the sequence of "N8-W-N21". The estimated barcode diversity of current batch of virus is ~45M. The titer of the virus is ~5x10^10 infectious particles/ml

 *price per 24 aliquot-- 4ul/aliquot

Provide Sindbis virus library containing millions of barcodes. The estimated barcode diversity of current batch of virus is ~19M. The titer of the virus is ~2x10^11 infectious particles/ml

 *price per 24 aliquot-- 4ul/aliquot

This test virus has the same vector sequence as the barcoded virus library, but it contains only one barcode. This virus could be used for practicing Sindbis virus injection. Each order contains 4 aliquots of virus (4ul/aliquot), which should be enough for at least 8 injections.

This service provides 200-300 um cryostat sections for manual dissection purpose from fresh frozen tissue. Extra care is taken during cryostat sectioning to avoid cross sample contamination. The sections are stored at -80oC before dissection.

We offer a preliminary Next Generation Sequencing (NGS) data processing service that includes the analysis of raw data and the generation of a Barcode Matrix, which provides information on the abundance of each barcoded neuron in each tissue sample.

Our pricing is based on a per sequencing run basis.

Starting from June 1st, 2023, our data processing service will undergo a significant update, resulting in a remarkable drop in the cost of data processing. Our new and improved pipeline is designed to be more efficient and faster, allowing us to deliver high-quality results at a much more competitive price point. This update represents our commitment to providing exceptional value to our users.

This package offers a comprehensive pilot study using both BARseq and MAPseq, designed to

help researchers generate preliminary data for grant applications or feasibility testing.

Included Services

1.Barcoded Sindbis Virus Library

o High-titer Sindbis virus: 10^10 ifu/mL

o Contains 40 million unique barcodes

o Supplied volume is sufficient to infect a 1 mm. brain area in at least 6 animals

o Developed and validated by the Zador lab for use in MAPseq and BARseq

2.Tissue Preparation

o Cryostat sectioning:

– 2 x 60 μm fixed sections for BARseq

– Up to 10 x 200 μm sections for MAPseq

o Manual dissection of up to 6 brain regions for MAPseq

o Dissection of 1 brain area, 2 sections for BARseq

3. BARseq Workflow

o One BARseq run on the selected region

o In situ sequencing:

– 7 cycles for gene sequencing

– 15 cycles for barcode sequencing

o Library preparation, rolony generation, and detection of barcode and 104 genes

4. MAPseq Workflow

o Library preparation for up to 6 dissected brain regions, including QC

o Pooled with other user sample for one high-throughput sequencing run

o Preliminary MAPseq data processing

Turnaround & Data Handling

• BARseq: The core provides the processing codes to users.

• MAPseq: Sequencing is pooled with other user samples; turnaround time depends on

demand.

Additional Notes

• Mouse brain injection and collection: Users are responsible for virus injection and

brain collection following the protocol provided by the core.

• Custom Probes: The package supports detection of 104 genes (Chen et al., Nature,

2024). Users may include additional probes at their own expense.

• Protocol Compliance: Deviations from the provided sample prep protocol may result in

extra charges.

Reference

Chen, X., Fischer, S., Rue, M.C.P. et al. Whole-cortex in situ sequencing reveals input-dependent area identity. Nature (2024). https://doi.org/10.1038/s41586-024-07221-6

Sequencing the MAPseq library by PE36 NextSeq

*Price per library

• 5B=1.25B paired-end reads

We are offering a free QC check on RNA quality and barcode amount for up to 10 mouse brain samples per lab. This service is available to labs from non-profit institutes between July 2024 and July 2027.

QC service includes:

• RNA extraction
• RIN check on a bioanalyzer
• RT-qPCR to examine barcode quantity

This free service is limited to 10 samples per lab.

RNA extraction, QC for RNA quality and injection quality (bioanalyzer and q-PCR), Reverse Transcription and PCR to make the library for next generation sequencing.

RNA extraction, QC for RNA quality and injection quality (bioanalyzer and q-PCR), Reverse Transcription and PCR to make the library for next generation sequencing.

RNA extraction, QC for RNA and injection quality (bioanalyzer and q-PCR), Reverse Transcription and PCR to make the library for next generation sequencing.

RNA extraction, QC for RNA and injection quality (bioanalyzer and q-PCR), Reverse Transcription and PCR to make the library for next generation sequencing

RNA extraction, QC for RNA and injection quality (bioanalyzer and q-PCR), Reverse Transcription and PCR to make the library for next generation sequencing.

Run bioanalyzer to check RNA quality and RT-qPCR to examine barcode amount. Price listed is per 10 samples. RNA extraction is not included in the price.

Extract RNA from mice brain samples with Trizol reagent.

Provide Sindbis virus library containing millions of barcodes with the sequence of "N8-W-N21". The estimated barcode diversity of current batch of virus is ~45M. The titer of the virus is ~5x10^10 infectious particles/ml

 *price per 24 aliquot-- 4ul/aliquot

Provide Sindbis virus library containing millions of barcodes. The estimated barcode diversity of current batch of virus is ~19M. The titer of the virus is ~2x10^11 infectious particles/ml

 *price per 24 aliquot-- 4ul/aliquot

This test virus has the same vector sequence as the barcoded virus library, but it contains only one barcode. This virus could be used for practicing Sindbis virus injection. Each order contains 4 aliquots of virus (4ul/aliquot), which should be enough for at least 8 injections.