AtaGenix Provides Custom CHALK10 and SD1 Protein Expression and Rabbit Polyclonal Antibody Services for Rice Chalkiness Research at China National Rice Research Institute

Study Context
Grain chalkiness in rice (Oryza sativa) negatively impacts grain quality, affecting texture, milling resilience, and consumer preference. A 2025 study published in Plant Communications (DOI: 10.1016/j.xplc.2025.101354) by researchers at the State Key Laboratory of Rice Biology and Breeding, China National Rice Research Institute, identified CHALK10, an F-box protein that negatively regulates grain chalkiness by mediating the ubiquitination and degradation of SEMIDWARF-1 (SD1) via the 26S proteasome pathway. The study revealed that CHALK10 interacts with SD1, influencing gibberellin (GA) levels and starch metabolism, thereby affecting endosperm development and chalkiness.

Client Requirements
The research team, led by Wei Zhou and colleagues at the State Key Laboratory of Rice Biology and Breeding, required high-purity recombinant CHALK10 and SD1 proteins for co-immunoprecipitation (co-IP) assays to validate their interaction in rice endosperm development. Additionally, they needed rabbit polyclonal antibodies against these proteins for immunoblotting in ubiquitination and degradation studies. AtaGenix Laboratories Co., Ltd. (Wuhan, China) was commissioned to provide custom protein expression, purification, and rabbit polyclonal antibody production services using an Escherichia coli expression system, ensuring compatibility with co-IP validation and downstream immunological assays.

Technical Challenges
1. Protein Expression: Achieving high-yield expression of functional CHALK10 and SD1 proteins in E. coli, which lacks eukaryotic post-translational modification systems.
2. Protein Purity: Ensuring high-purity proteins for sensitive co-IP assays to confirm protein-protein interactions.
3. Antibody Specificity: Producing rabbit polyclonal antibodies with high specificity and sensitivity for detecting CHALK10 and SD1 in complex rice protein extracts.

Custom Protein and Antibody Solution
AtaGenix collaborated with the State Key Laboratory of Rice Biology and Breeding to deliver custom protein expression and antibody production services for their rice chalkiness study published in Plant Communications (DOI: 10.1016/j.xplc.2025.101354). Our solution included:

1. Protein Expression and Purification:
- Expressed CHALK10 and SD1 proteins in E. coli Rosetta (DE3) cells using pMAL-c2X (MBP-tagged CHALK10) and pGEX-6P-1 (GST-tagged SD1) vectors for high-yield production.
- Purified recombinant proteins to high purity using glutathione and amylose affinity chromatography, ensuring suitability for co-IP and in vitro ubiquitination assays.

2. Rabbit Polyclonal Antibody Production:
- Immunized rabbits with purified CHALK10 and SD1 proteins to generate high-titer polyclonal antibodies.
- Validated antibody specificity through Western blotting, confirming strong and specific detection of CHALK10 and SD1 in rice protoplast extracts.

3. Validation and Support:
- Supported co-IP assays to confirm CHALK10-SD1 interaction, enabling the research team to validate the role of CHALK10 in SD1 degradation via the 26S proteasome pathway.
- Provided technical assistance for optimizing antibody performance in immunoblotting, facilitating accurate detection of ubiquitinated SD1 in in vivo assays.

The high-purity CHALK10 and SD1 proteins, along with specific rabbit polyclonal antibodies, enabled the research team to confirm the CHALK10-SD1 interaction and its role in regulating rice grain chalkiness through GA-mediated starch metabolism. Validated through co-IP, these reagents advanced the understanding of molecular mechanisms underlying rice grain quality.

For more information, visit www.atagenix.com. AtaGenix provides one-stop customized support from antibody/protein design to functional validation, covering custom antibodies, protein expression, phage display, and multi-platform validation.