Researchers have cloned and characterized a cDNA sequence of the ABCD4 gene from Ujimqin sheep, adding a new molecular resource for livestock genetics. ABCD4 encodes an ATP-binding cassette transporter involved in intracellular cobalamin trafficking, and the gene has also been identified as a candidate linked to variation in vertebral number in livestock. Until now, molecular information and experimental tools for the ovine version of the gene have remained limited, which is what the new study set out to address.
The team, working with liver tissue from Ujimqin sheep, used rapid amplification of cDNA ends, a technique known as RACE, to recover the full transcript sequence.
The resulting sequence measured 1,874 base pairs in length. Within it, the researchers identified a predicted open reading frame of 1,209 base pairs, which encodes a polypeptide of 402 amino acids. Sequence and bioinformatic analyses showed that the RACE-derived sequence retained conserved features characteristic of the ABC transporter family, although the predicted protein it encodes is shorter than the canonical ABCD4 proteins described in other mammals.
Beyond the sequence itself, the study produced two practical laboratory resources. The researchers built a recombinant protein expression system for ABCD4 and generated a polyclonal antibody against the protein, tools that can support future work on where the gene is expressed, where the protein localizes within cells, and what it does biologically in sheep.
To produce the recombinant antigen, the ABCD4 expression construct was introduced into Escherichia coli BL21 (DE3), a standard bacterial strain used for protein production. The researchers tested induction conditions and found that higher recombinant protein expression was achieved when cultures were induced with 0.5 millimolar IPTG at 37 degrees Celsius for four hours.
The recombinant protein was predominantly recovered from the insoluble fraction of the bacterial cells. The team nevertheless purified the protein using Ni-NTA affinity chromatography.
The purified recombinant protein was then used to immunize New Zealand White rabbits solely for the purpose of producing polyclonal antiserum. The resulting antisera showed strong immunoreactivity against the recombinant antigen.
The work is descriptive and methodological rather than functional. The authors did not report experiments testing what ABCD4 does in living sheep, and the connection to vertebral-number variation remains a candidate-gene association noted in the broader literature rather than something demonstrated in this study. The predicted protein length discrepancy with canonical mammalian ABCD4 also suggests that the RACE-derived sequence may not capture the full canonical coding region, a point the researchers acknowledge in describing the product as shorter than its mammalian counterparts.
Even so, the study fills a practical gap.
The authors position the work explicitly as a foundation: a RACE-derived cDNA sequence, a working prokaryotic expression system, and an antibody resource that may support future investigations of ABCD4 expression, localization, and biological function in sheep.
Subject of Research: Cloning and molecular characterization of the ovine ABCD4 transporter gene and development of recombinant protein and antibody resources in Ujimqin sheep
Article Title: Cloning and molecular characterization of an ovine ABCD4 cDNA sequence, recombinant protein expression, and polyclonal antibody generation in Ujimqin sheep
Article References: Zhang, Y., Zhang, J., Zhao, J., Jiang, L., Zhao, Q., Wu, J., Jian, R., Yuan, G., Zhao, S., Cao, G., Zhang, Y., Bao, F., & Liu, J. (2026). Cloning and molecular characterization of an ovine ABCD4 cDNA sequence, recombinant protein expression, and polyclonal antibody generation in Ujimqin sheep. Frontiers in Veterinary Science, 13, Article 1869570. https://doi.org/10.3389/fvets.2026.1869570
Image Credits: AI Generated
DOI: 10.3389/fvets.2026.1869570
Keywords: ABCD4, Ujimqin sheep, cDNA cloning, RACE, ATP-binding cassette transporter, cobalamin trafficking, recombinant protein expression, E. coli, Ni-NTA purification, polyclonal antibody, livestock genomics, vertebral number
News Source: Juliet Wilcox. (October 28, 2026). Scientists Clone Ovine ABCD4 Gene and Build Antibody Resource for Sheep Research. Scienmag.
