Takara SMART-Seq Mouse BCR (with UMIs), 96 Rxns

SMART-Seq Mouse BCR (with UMIs) enables users to analyze B-cell receptor (BCR) repertoires from 10 ng–1 µg high-integrity (RIN>7) bulk total RNA samples from spleen, bone marrow, or peripheral blood mononuclear cells (PBMCs) or RNA extracted from mouse whole blood (RNA input ≥100 ng) or lymph nodes. The kit can be used to generate data for both heavy (IgG, IgM, IgA, IgD, and IgE) and light chains (IgK, IgL) of mouse B-cell receptors.

Schematic of technology and workflow for SMART-Seq Muman BCR (with UMIs).Input RNA or cells are oligo-dT primed using the dT Primer (dark blue). Following oligo dT-priming, SmartScribe Reverse Transcriptase performs first strand cDNA synthesis on input RNA or cells and adds non-templated nucleotides to the 5′ end of each cDNA molecule (XXXXX). Upon reaching the 5′ end of the RNA template, the SMART UMI Oligo anneals to the non-templated nucleotides (XXXXX), incorporating the UMI (yellow) and partial Illumina adapter (light green) complementary to the mBCR PCR1 Universal Forward primer. Following reverse transcription, semi-nested PCR is performed to amplify BCR cDNAs. In PCR1, the mBCR PCR1 Universal Forward primer anneals to the complementary sequence carried by the SMART UMI Oligo (light green), incorporating Illumina Read2 sequence (dark green). The mBCR PCR1 Reverse primer (orange) anneals to sequences in the constant regions of BCR cDNAs to amplify the entire BCR V(D)J region. During the PCR2, the nested mBCR LC (light chain) or mBCR HC (heavy chain) PCR2 Reverse primers anneal to sequences in BCR constant regions that are internal to the sequences bound by the mBCR PCR1 Reverse primer and add the Illumina Read 1 sequence (light purple). In the same PCR2 reaction, Unique Dual Index Kit primers anneal to sequence added by mBCR PCR1 Universal Forward primer or the mBCR LC or HC PCR2 Reverse primers to add Illumina P7-i7 and P5-i5 index sequences (dark blue). The result is a sequencing-ready library that contains the entire BCR light chain and/or heavy chain variable regions with a small portion of the constant region.

Panel A.Detection of all BCR isotypes for both heavy chain (IgA/D/g/m, except IgE due to its very low expression in normal mouse samples) and light chain (IgK/L) by SMART-Seq Mouse BCR (with UMIs) in 10, 100, and 1,000 ng input of mouse spleen RNA. Comparison across various RNA inputs shows a consistent increase in clonotype counts with higher input.Panel B.Sensitivity of SMART-Seq Mouse BCR (with UMIs) ensures detection of more heavy-chain and light-chain clonotypes with more input RNA. bar graphs illustrate both heavy-chain and light-chain clonotype counts consistently increase with a higher amount of input RNA.

Required Products

The Unique Dual Index (UDI) Kit (1–96) contains 96 pairs of unique dual-indexed PCR primers that can be used to generate up to 96 Illumina-compatible sequencing libraries and are recommended with several of Takara Bio’s RNA-seq and DNA-seq kits.

The Unique Dual Index (UDI) Kit (97–192) contains 96 pairs of unique dual-indexed PCR primers that can be used to generate up to 96 Illumina-compatible sequencing libraries and are recommended with several of Takara Bio’s RNA-seq and DNA-seq kits.

The Unique Dual Index (UDI) Kit (193–288) contains 96 pairs of unique dual-indexed PCR primers that can be used to generate up to 96 Illumina-compatible sequencing libraries and are recommended with several of Takara Bio’s RNA-seq and DNA-seq kits.

The Unique Dual Index (UDI) Kit (289–384) contains 96 pairs of unique dual-indexed PCR primers that can be used to generate up to 96 Illumina-compatible sequencing libraries and are recommended with several of Takara Bio’s RNA-seq and DNA-seq kits.

The Unique Dual Index (UDI) Kit (1–24) contains 24 pairs of unique dual-indexed PCR primers, which are a subset of the Unique Dual Index Kit (1–96), Cat. No. 634752. They can be used to generate up to 24 Illumina-compatible sequencing libraries and are recommended with several of Takara Bio’s RNA-seq and DNA-seq kits.

NucleoMag NGS Clean-up and Size Select employs scalable, automation-friendly magnetic bead technology to enable efficient cleanup of DNA or RNA fragments. The flexible protocol enables processing of 50–150 µl input volumes and allows for enrichment of fragments ranging in size from 150–800 bp. NucleoMag NGS Clean-up and Size Select consists of paramagnetic beads suspended in binding buffer that selectively bind DNA and RNA fragments based on the volume ratio of bead suspension to sample. After magnetic separation and removal of supernatant, the beads are washed with ethanol followed by a short drying step. Following elution in low-salt buffer or water, the highly purified DNA is free of contaminants such as nucleotides, primers, adapters, adapter dimers, enzymes, buffer additives, or salts, and can be used directly in downstream applications. NucleoMag NGS Clean-up and Size Select provides comparable performance and employs the same dilution ratios for size selection as magnetic bead-based solutions from other providers, such that it can be incorporated in existing workflows without the need for protocol adjustment or optimization.

Cat. # 744970.5 consists of a bottle containing 5 mL of bead suspension.

NucleoMag NGS Clean-up and Size Select employs scalable, automation-friendly magnetic bead technology to enable efficient cleanup of DNA or RNA fragments. The flexible protocol enables processing of 50–150 µl input volumes and allows for enrichment of fragments ranging in size from 150–800 bp. NucleoMag NGS Clean-up and Size Select consists of paramagnetic beads suspended in binding buffer that selectively bind DNA and RNA fragments based on the volume ratio of bead suspension to sample. After magnetic separation and removal of supernatant, the beads are washed with ethanol followed by a short drying step. Following elution in low-salt buffer or water, the highly purified DNA is free of contaminants such as nucleotides, primers, adapters, adapter dimers, enzymes, buffer additives, or salts, and can be used directly in downstream applications. NucleoMag NGS Clean-up and Size Select provides comparable performance and employs the same dilution ratios for size selection as magnetic bead-based solutions from other providers, such that it can be incorporated in existing workflows without the need for protocol adjustment or optimization.

Cat. # 744970.50 consists of a bottle containing 50 mL of bead suspension.

NucleoMag NGS Clean-up and Size Select employs scalable, automation-friendly magnetic bead technology to enable efficient cleanup of DNA or RNA fragments. The flexible protocol enables processing of 50–150 µl input volumes and allows for enrichment of fragments ranging in size from 150–800 bp. NucleoMag NGS Clean-up and Size Select consists of paramagnetic beads suspended in binding buffer that selectively bind DNA and RNA fragments based on the volume ratio of bead suspension to sample. After magnetic separation and removal of supernatant, the beads are washed with ethanol followed by a short drying step. Following elution in low-salt buffer or water, the highly purified DNA is free of contaminants such as nucleotides, primers, adapters, adapter dimers, enzymes, buffer additives, or salts, and can be used directly in downstream applications. NucleoMag NGS Clean-up and Size Select provides comparable performance and employs the same dilution ratios for size selection as magnetic bead-based solutions from other providers, such that it can be incorporated in existing workflows without the need for protocol adjustment or optimization.

Cat. # 744970.500 consists of a bottle containing 500 mL of bead suspension.