# Rapid Identification of the SNP Mutation in the ABCD4 Gene and Its Association with Multi-Vertebrae Phenotypes in Ujimqin Sheep Using TaqMan-MGB Technology

## TL;DR

A new, affordable genetic test was developed to identify Ujimqin sheep with extra vertebrae, a trait linked to higher meat yield, using a specific gene mutation.

## Contribution

A TaqMan-MGB genotyping method was developed for rapid and accurate detection of a key SNP in Ujimqin sheep associated with multi-vertebrae traits.

## Key findings

- The TaqMan-MGB test showed 100% agreement with Sanger sequencing and 91.4% accuracy in detecting multi-vertebrae sheep.
- The test is simpler, faster, and cheaper than traditional imaging methods like DR.
- The SNP ABCD4 Chr7:89393414 C > T is strongly associated with multi-vertebrae phenotypes in Ujimqin sheep.

## Abstract

This study developed a fast, cost-effective genetic test to identify Ujimqin sheep with extra vertebrae, a trait linked to higher meat yield. Researchers targeted a specific gene change (SNP: ABCD4, Chr7:89393414 C > T) known to cause extra vertebrae. We designed a TaqMan-MGB test using fluorescent probes to detect this SNP in blood DNA. Validated on 152 sheep, the test showed 100% agreement with gold-standard DNA sequencing (Sanger method). Compared to DR imaging (83.6% overall agreement), it accurately detected 91.4% of multi-vertebrae sheep (127/139 animals). The test is simple, high-throughput, and significantly cheaper than DR imaging. This tool enables early, efficient selection of breeding sheep with the desirable multi-vertebrae trait, accelerating genetic improvement programs for Ujimqin sheep farmers.

Ujimqin sheep, known for its distinctive multi-vertebrae phenotypes (T13L7, T14L6, and T14L7) and economic value, has garnered significant attention. However, conventional phenotypic detection methods suffer from low efficiency and high costs. In this study, based on a key SNP locus (ABCD4 gene, Chr7:89393414, C > T) identified through a genome-wide association study (GWAS), a TaqMan-MGB (minor groove binder) genotyping system was developed. the objective was to establish a high-throughput and efficient molecular marker-assisted selection (MAS) tool. Specific primers and dual fluorescent probes were designed to optimize the reaction system. Standard plasmids were adopted to validate genotyping accuracy. A total of 152 Ujimqin sheep were subjected to TaqMan-MGB genotyping, digital radiography (DR) imaging, and Sanger sequencing. the results showed complete concordance between TaqMan-MGB and Sanger sequencing, with an overall agreement rate of 83.6% with DR imaging. For individuals with T/T genotypes (127/139), the detection accuracy reached 91.4%. This method demonstrated high specificity, simplicity, and cost-efficiency, significantly reducing the time and financial burden associated with traditional imaging-based approaches. the findings indicate that the TaqMan-MGB technique can accurately identify the T/T genotype at the SNP site and its strong association with the multi-vertebrae phenotypes, offering an effective and reliable tool for molecular breeding of Ujimqin sheep.