Nanobody-Tn5 is a recombinant Tn5 transposase fused with nanobody which can specific bind with primary antibodies from two different species (mouse and rabbit). Components Component No. Component name 12595ES01 12595ES03 12595-A Anti-mouse Tn5 10 μL 100 μL 12595-B Anti-rabbit Tn5 10 μL 100 μL 12595-C Assemble buffer 10 μL 100 μL Specifications Cat.No. 12595ES01 /12595ES03 Unit Size 10 μL / 100 μL Protein Concentration Anti-mouse Tn5: 0.55 mg/mL Anti-rabbit Tn5: 0.65 mg/mL Protein Molecular Weight 67 kDa Transposase Activation Conditions 37℃ 1 h 1 × Annealing Buffer( 10 mM Tris-HCl, 50 mM NaCl, pH 7.5–8.0(±0.1 mM EDTA) Figures H3K4me1 H3K4me3 H3K9me3 H3K27ac H3K27me3 H3K27me3 Control: Conventional pA/G-Tn5, Cut&Tag Multi Cut&Tag: Nanobody Tn5 Instruction Take high-throughput sequencing library preparation as an example, be sure to read the instructions carefully before use. 1. Adapter preparation 1) Reference oligo name and sequence of Illumina platform: Oligo A-1：5'-AATGATACGGCGACCACCGAGATCTACACATAAGGCTCGTCGGCAGCGTCAGATGTGTATAAGAGACAG-3' Oligo A-2：5'-AATGATACGGCGACCACCGAGATCTACACATATCCTTCGTCGGCAGCGTCAGATGTGTATAAGAGACAG-3' Oligo A-3：5'-TCGTCGGCAGCGTCAGATGTGTATAAGAGACAG-3′ Oligo B：5'-GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAG-3′Oligo C：5'-phos-CTGTCTCTTATACACATCT-3' 2) Dissolve the Oligo A, Oligo B, and Oligo C to 200 μM. 3) The following reaction systems are formulated respectively: Components Reaction 1 Oligo B （200 μM） 10 μL Oligo C （200 μM） 10 μL total 20 μL Components Reaction 2 Oligo A-1 or A-2 or A-3（200 μM） 10 μL Oligo C （200 μM） 10 μL total 20 μL 4) Mix reaction 1 and 2 together, and briefly centrifuge to bring the solution back to the bottom of the tube. Place in the PCR instrument, set up the following program and run it: hot lid at 105°C,and then heat at 94℃ for 2 min. After the time was up, the reaction procedure was stopped without opening the hot lid.The PCR tube was naturally cooled in the PCR instrument for 2 h, and then placed at 4℃ for 5 min. 5) After the reaction, mix the equal volumes of Reaction 1 and Reaction 2. Named ad Adapter Mix and stored from – 25 to – 15℃. 2. For transposase generation 1) To configure the following reaction system: Components volume (μL) Anti-mouse Tn5 or Anti-rabbit Tn5 10 Adapter mix (50 μM) 0.5 Assemble Buffer 0.5 2) Reaction conditions: use the pipette to gently blow and mix well. Perform the reaction at 25℃ for 1 h (hot lid off). The products of reaction can be directly applied to the library preparation or stored from -25 to -15℃. Notes 1. Transposase is temperature-sensitive, so it is recommended to complete the adapter embedding as soon as possible. The generated transposon can be stored at-25~ -15℃, with a validity period of 1 year. 2. This product is used for scientific research purposes only. 3. For your safety and health, please wear laboratory clothes and wear disposable gloves.
