Anti-DNA-RNA hybrid mAbs [S9.6](MABL-1414)

MABL-1414

ABA, CHIP, ChIP-seq, DB, EMSA, FISH, ICC, IP, PLA, SPR, IF

Engineer antibody

Species independent

S9.6

Recombinant Antibody

This antibody demonstrates high specificity and affinity for DNA/RNA hybrids and other A-form nucleic acid hybrids, making it an effective tool for detecting R-loops. It does not cross-react with single- stranded DNA, double-stranded DNA, RNA, or ribosomal RNA. In the original study by Boguslawski et al. (1986), the antibody was shown to bind DNA–RNA heteropolymers and poly(I)–poly(dC) with equal affinity, though 100-fold higher concentrations of poly(A)–poly(dT) were required for comparable binding. DNA–RNA hybrids naturally occur in eukaryotic cells, particularly at sites of high transcriptional activity. These non- canonical nucleic acid structures have transcriptional regulatory functions and are associated with genomic instability. The presence of DNA–RNA hybrids can predispose specific loci to chromosomal breakage. When formed, these hybrids create a double-stranded A/B intermediate conformation, exposing the displaced DNA strand as a target for single-stranded nucleic acid-binding proteins. Additionally, DNA–RNA hybrids are resistant to the action of DNA methyltransferases. The formation of these hybrids has also been linked to several neurological diseases. Mutations in the DNA–RNA helicase senataxin (SETX) are associated with the dominant juvenile form of amyotrophic lateral sclerosis type 4 and a recessive form of ataxia with oculomotor apraxia type 2, highlighting their clinical relevance.

DNA:RNA Duplex; DNA-RNA Duplex; DNA/RNA Duplex; DNA-RNA Hybrid; DNA/RNA Hybrid; DNA:RNA Hybrid; RNA-DNA Duplex; RNA/DNA Duplex; RNA:DNA Duplex; RNA-DNA Hybrid; RNA/DNA Hybrid; RNA:DNA Hybrid

This antibody was generated in BALB/c mouse against S9.6 ΦX174 bacteriophage-derived synthetic DNA/RNA antigen according to the protocol by Boguslawski et al. (J. Immunol Methods. 1986).

DNA–RNA hybrids naturally occur in eukaryotic cells, with their levels increasing in regions of high transcriptional activity, such as during transcription initiation, repression, and elongation. These hybrids play a crucial role in influencing genomic instability, making the anti-DNA–RNA hybrid (S9.6) antibody an invaluable tool for studying R-loops and hybrid-induced lesions during DNA replication and other cellular processes. This antibody also excels in recognizing RNA–DNA hybridization in microarray studies. Key applications of the S9.6 antibody include affinity binding assays (ABA), chromatin immunoprecipitation (ChIP), chromatin immunoprecipitation-sequencing (ChIP-seq), dot blot (DB), fluorescent in situ hybridization (FISH), electrophoretic mobility shift assays (EMSA), surface plasmon resonance (SPR), immunofluorescence (IF), immunocytochemistry (ICC), and immunoprecipitation (IP). For example, in ABA, this antibody binds DNA–RNA heteropolymers and poly(I)–poly(dC) equally, but requires 100-fold higher levels of poly(A)–poly(dT) to achieve similar binding levels (Boguslawski et al., 1986; PMID: 2422282). In a ChIP analysis, a representative lot detected elevated DNA–RNA hybrid levels at four actively transcribed genes following shRNA-mediated knockdown of BRCA1 or BRCA2, but not PCID2 or RAD51 in HeLa cells (Bhatia et al., 2014; PMID: 24896180). Similarly, in ChIP-seq, the antibody detected genome-wide distribution of DNA–RNA hybrids in budding yeast (El Hage et al., 2014; PMID: 25357144). In ICC and IP analyses, the antibody immunoprecipitated in vitro transcribed R-loop substrates (DNA–RNA hybrids) but not double- stranded DNA (dsDNA) (Ginno et al., 2012; PMID: 22387027). Additionally, this antibody has been used for IF and proximity ligation assays (PLA) to detect DNA–RNA hybrids. In IF, it revealed DNA–RNA hybrid accumulation in HeLa cells during S/G2/M phases following ATAD5 depletion. In PLA, it assessed the proximity between DNA–RNA hybrids and PCNA (Kim et al., 2020; PMID: 32542338). Moreover, the Fab version of this antibody demonstrated binding to a 13-bp DNA/RNA hybrid, which was further studied via X- ray crystallography to examine its binding structure (Bou-Nader et al., 2022; PMID" 32542338). Moreover, the Fab version of this antibody demonstrated binding to a 13-bp DNA/RNA hybrid, which was further studied via X- ray crystallography to examine its binding structure (Bou-Nader et al., 2022; PMID: 35347133).

Boguslawski et al. Characterization of monoclonal antibody to DNA.RNA and its application to immunodetection of hybrids. J Immunol Methods. 1986 May 1;89(1):123-30. PMID:2422282

The original publication describes the generation of this antibody, its characterization, and its application to the immunodetection of DNA-RNA heteropolymer hybrids.

100 μg Purified antibody.

1 mg/ml.

Protein A affinity purified

PBS with 0.02% Proclin 300.

Store at 4⁰C for up to 3 months. For longer storage, aliquot and store at - 20⁰C.