GeneRulor Nanobody-Tn5 Transposase

1. Product Overview

GeneRulor Nanobody-Tn5 Transposase is a fusion protein engineered via protein engineering technology, which conjugates a specific nanobody with a hyperactive Tn5 transposase and can efficiently recognize the IgG Fc fragment of mouse- or rabbit-derived primary antibodies. Its nanobody module is only 15 kDa, with a volume reduced by 90% compared with traditional antibodies, and possesses high affinity (picomolar-level binding capacity) and excellent thermal stability. This innovative design allows the Tn5 transposase to directly target the binding sites of primary antibodies, insert sequencing adapters with unique indexes while cleaving chromatin, and eliminate the step of secondary antibody incubation, significantly simplifying the experimental workflow. By preloading different Barcode adapters, parallel detection of multiple targets (e.g., histone modifications such as H3K27ac/H3K27me3) can be achieved in a single experiment with a markedly improved signal-to-noise ratio. The technology has been successfully applied to single-cell multimodal epigenetic analysis, supporting simultaneous resolution of chromatin states and cell surface proteins, and providing a high-resolution tool for research in immunology, neural development and disease mechanisms.

2. Product Features

(1) Ultra-high affinity and specificity: The affinity of the nanobody for the target IgG is superior to traditional Protein-A/G, enabling high-precision localization of chromatin markers.

(2) Excellent stability: Maintains high activity over a wide temperature range and under different pH conditions, ensuring the stability and reliability of experimental results and suitability for various complex samples.

(3) Flexible multiplex detection: With species-specific nanobodies, simultaneous analysis of multiple histone modifications or transcription factor binding sites can be realized in a single experiment.

(4) Small size and low background interference: The nanobody has a molecular weight of only 15 kDa, with excellent tissue penetration ability and extremely low non-specific binding, leading to a significant increase in the signal-to-noise ratio.

(5) Ready-to-use single-cell compatible design: Preloadable with Barcode adapters, it can be directly applied to single-cell platforms such as 10x Genomics with a simple workflow and sensitivity covering the single-cell level.

3. Applications

NTT-seq Technology: GeneRulor Nanobody-Transposase is basically applied to cutting-edge epigenetic research, especially achieving multimodal chromatin analysis at single-cell resolution via NTT-seq. Its working principle is based on the specific binding of nanobodies to primary antibodies of different species or IgG subtypes (e.g., anti-mouse/rabbit IgG), guiding the Tn5 transposase to target histone modifications (e.g., H3K27ac, H3K27me3) or transcription factor binding sites, and completing in situ DNA fragmentation and barcode adapter insertion. This method features high sensitivity (capable of detecting as low as the single-cell level), low background interference and multiplex advantages, supporting the simultaneous resolution of multiple chromatin markers.

Figure 1. Experimental flow chart of CUT&Tag using GeneRulor Nanobody-Transposase

References

[1] Kaya-Okur HS, Wu SJ, Codomo CA, Pledger ES, Bryson TD, Henikoff JG, Ahmad K, Henikoff S. CUT&Tag for efficient epigenomic profiling of small samples and single cells. Nat Commun. 2019 Apr 29;10(1):1930. doi: 10.1038/s41467-019-09982-5.

[2] Farzad N, Enninful A, Bao S, Zhang D, Deng Y, Fan R. Spatially resolved epigenome sequencing via Tn5 transposition and deterministic DNA barcoding in tissue. Nat Protoc. 2024 Nov;19(11):3389-3425. doi: 10.1038/s41596-024-01013-y.

[3] Stuart T, Hao S, Zhang B, Mekerishvili L, Landau DA, Maniatis S, Satija R, Raimondi I. Nanobody-tethered transposition enables multifactorial chromatin profiling at single-cell resolution. Nat Biotechnol. 2023 Jun;41(6):806-812. doi: 10.1038/s41587-022-01588-5.

[4] Bartosovic M, Castelo-Branco G. Multimodal chromatin profiling using nanobody-based single-cell CUT&Tag. Nat Biotechnol. 2023 Jun;41(6):794-805. doi: 10.1038/s41587-022-01535-4.