Recombinant Human FANCI Protein: Research Guidelines

Recombinant Human FANCI Protein is a key component of the Fanconi anemia DNA repair pathway, functioning as a monoubiquitination substrate and chromatin-recruitment factor essential for resolving DNA interstrand crosslinks in research applications.

Product Name

Recombinant Human FANCI Protein

SKU

rp377702

Gene / Protein Target

FANCI (Fanconi Anemia Complementation Group I)

Biological Function

Forms the FANCD2-FANCI (ID2) complex; required for monoubiquitination and nuclear foci formation during DNA interstrand crosslink repair

Primary Research Applications

DNA repair assays, Fanconi anemia pathway studies, protein-protein interaction studies, biochemical reconstitution

Recommended Storage

-80 °C for long-term storage; avoid repeated freeze-thaw cycles

Product Overview and Biological Background

Recombinant Human FANCI Protein (SKU: rp377702) is a recombinant form of the human Fanconi Anemia Complementation Group I (FANCI) protein. FANCI is an essential component of the Fanconi anemia (FA) DNA damage response pathway, which is the primary cellular mechanism for repairing DNA interstrand crosslinks (ICLs). ICLs are highly cytotoxic lesions that covalently link both strands of the DNA double helix, blocking replication and transcription.

FANCI forms a stable heterodimeric complex with FANCD2, known as the ID2 complex. This complex is a central node of the FA pathway: upon DNA damage or replication stress, both FANCI and FANCD2 are monoubiquitinated by the FA core ubiquitin E3 ligase complex. Monoubiquitination triggers localization of the ID2 complex to chromatin at sites of ICL damage, where it coordinates downstream repair activities including nucleolytic incision, translesion synthesis, and homologous recombination.

Mutations in the FANCI gene cause Fanconi anemia subtype I (FA-I), a rare inherited disorder characterized by bone marrow failure, developmental anomalies, and a markedly elevated risk of malignancy. The recombinant protein offered by Aladdin Scientific supports mechanistic and translational research into this critical pathway.

Reconstitution and Handling Protocol

Reconstitution: Upon receipt, briefly centrifuge the vial at low speed (e.g., 2,000 × g for 30 seconds) to collect lyophilized material at the bottom before opening. Reconstitute the protein in an appropriate buffer such as sterile phosphate-buffered saline (PBS, pH 7.4) containing a carrier protein (e.g., 0.1% BSA) or a buffer recommended for your downstream assay. Add the reconstitution buffer slowly and gently mix by pipetting; do not vortex.

Working concentration: Determine empirically for each experimental system. As a general starting point for biochemical assays, prepare a stock solution at the concentration indicated on the product datasheet and dilute as required.

Handling precautions:

• Work on ice at all times to preserve protein activity.
• Prepare single-use aliquots immediately after reconstitution to minimize freeze-thaw cycles.
• Use low-binding microtubes to reduce protein adsorption losses.
• Avoid introducing air bubbles during pipetting.

Quality check: Confirm integrity by SDS-PAGE or western blot before use in critical assays. A single band at the expected molecular weight confirms product integrity.

Storage Conditions and Stability

Short-term storage (up to 2 weeks): Store reconstituted protein at 4 °C. Addition of a carrier protein such as 0.1% BSA can help stabilize the protein during short-term refrigerated storage.

Long-term storage: Aliquot the reconstituted protein into single-use volumes and store at -80 °C. Properly stored aliquots are expected to remain stable for up to 12 months from the date of receipt; however, verify activity prior to use in time-sensitive experiments.

Avoid:

• Repeated freeze-thaw cycles, which can cause protein aggregation and loss of biological activity.
• Storage at -20 °C in frost-free freezers, which undergo temperature cycling.
• Prolonged exposure to room temperature.

Note: Specific stability data, expiry date, and exact storage buffer composition are provided on the product datasheet supplied with each lot. Always refer to the lot-specific datasheet for authoritative information.

Experimental Applications and Usage Recommendations

Recombinant Human FANCI Protein is suitable for a range of in vitro and cell-free biochemical studies focused on the Fanconi anemia pathway and DNA repair biology:

• ID2 complex reconstitution: FANCI can be combined with recombinant FANCD2 protein to reconstitute the ID2 heterodimer for structural and functional studies, including monoubiquitination assays.
• Protein-protein interaction studies: Use in pull-down assays, co-immunoprecipitation controls, or surface plasmon resonance (SPR) experiments to map interaction interfaces with FA core complex components or downstream repair factors.
• DNA-binding assays: FANCI harbors DNA-binding activity and can be used in electrophoretic mobility shift assays (EMSA) or fluorescence polarization assays to characterize its interaction with various DNA substrates, including ICL-containing duplexes.
• Ubiquitination assays: As a substrate for the FA core E3 ligase complex, recombinant FANCI can be used in cell-free ubiquitination assays to study pathway activation and regulation.
• Antibody validation: Recombinant FANCI protein serves as a positive control antigen for validating FANCI-specific antibodies by western blot or ELISA.

Optimization of protein concentration, buffer conditions, and assay parameters is recommended for each specific application. Consult the product datasheet for any application-specific guidance provided by Aladdin Scientific.

Quality, Safety, and Regulatory Notes

Quality assurance: Each lot of Recombinant Human FANCI Protein undergoes quality control testing prior to release. Specific QC data including purity assessment (typically by SDS-PAGE and Coomassie staining), endotoxin level, and biological activity results are reported on the lot-specific Certificate of Analysis (CoA). Request the CoA from Aladdin Scientific at the time of ordering or upon receipt.

Purity and endotoxin: For sensitive cell-based assays, verify that endotoxin levels meet requirements for your experimental system. If endotoxin contamination is a concern, consult the CoA or contact Aladdin Scientific technical support.

Safety: Handle recombinant proteins according to standard laboratory biosafety practices for BSL-1 materials. Wear appropriate personal protective equipment (gloves, lab coat, eye protection). The protein is not classified as hazardous under standard laboratory conditions; however, consult the Safety Data Sheet (SDS) provided for complete safety information.

Export and regulatory compliance: This product is intended for research use only (RUO) and is not approved for diagnostic or therapeutic applications in humans or animals. Users are responsible for ensuring compliance with all applicable local, national, and international regulations governing the import, export, and use of biological research materials.

Frequently asked questions

What buffer should I use to reconstitute Recombinant Human FANCI Protein?

Sterile PBS (pH 7.4) supplemented with 0.1% BSA is a commonly recommended reconstitution buffer for recombinant proteins to maintain solubility and activity. Always refer to the lot-specific datasheet supplied with your product for the manufacturer's recommended reconstitution buffer and volume, as these may be optimized for this specific protein.

How should I store Recombinant Human FANCI Protein to preserve its activity?

For long-term storage, aliquot the reconstituted protein into single-use volumes and store at -80 °C to avoid repeated freeze-thaw cycles, which can lead to aggregation and activity loss. For short-term use within two weeks, storage at 4 °C with a carrier protein such as BSA is acceptable. Avoid frost-free -20 °C freezers due to their temperature cycling.

What is the role of FANCI in the Fanconi anemia pathway?

FANCI partners with FANCD2 to form the ID2 heterodimeric complex, which is the central effector of the Fanconi anemia DNA interstrand crosslink repair pathway. Upon DNA damage, both FANCI and FANCD2 are monoubiquitinated by the FA core E3 ligase complex, triggering their recruitment to chromatin at ICL sites where they orchestrate downstream repair processes including nucleolytic incision, translesion synthesis, and homologous recombination.

Can Recombinant Human FANCI Protein be used for antibody validation?

Yes. Recombinant Human FANCI Protein (rp377702) is suitable as a positive control antigen for validating anti-FANCI antibodies by western blot or ELISA, providing a defined, pure reference standard for confirming antibody specificity and sensitivity.

Is Recombinant Human FANCI Protein suitable for cell-based or in vivo studies?

This product is designed and validated primarily for in vitro biochemical and cell-free assays. It is supplied for research use only (RUO) and is not approved for diagnostic or therapeutic applications. Suitability for cell-based assays depends on endotoxin levels and specific assay requirements; consult the Certificate of Analysis and contact Aladdin Scientific technical support for guidance.

References

1. Smogorzewska A, et al. Identification of the FANCI gene, the Fanconi anemia complementation group I gene. Cell. 2007;129(2):289-301. doi:10.1016/j.cell.2007.03.032
2. Sims AE, et al. FANCI is a second monoubiquitinated member of the Fanconi anemia pathway. Nature Structural & Molecular Biology. 2007;14(6):564-567. doi:10.1038/nsmb1252
3. Moldovan GL, D'Andrea AD. How the Fanconi anemia pathway guards the genome. Annual Review of Genetics. 2009;43:223-249. doi:10.1146/annurev-genet-102108-134222
4. Liang CC, et al. The FANCD2-FANCI complex is recruited to DNA interstrand crosslinks before monoubiquitination of FANCD2. Nature Communications. 2016;7:12124. doi:10.1038/ncomms12124

Products are supplied for research and development use only. Not for use in humans, animals, diagnosis, or therapy.