Evaluation of aptamers binding ability towards bovine X-specific sperm

Area of Science:

• Biotechnology
• Molecular Biology
• Animal Science

Background:

• Semen sexing is crucial for livestock breeding, with current research focusing on DNA aptamers for X-sorted sperm selection.
• DNA aptamers (Synthetic single-stranded DNA) offer a potential tool for sperm sex determination.

Purpose of the Study:

• To select and characterize DNA aptamers with high specificity for X-sorted sperm.
• To establish a proof of concept for aptamer-based semen sexing technology.

Main Methods:

• Cell-SELEX (Systematic Evolution of Ligands by Exponential Enrichment) was employed over four rounds.
• Aptamer specificity was assessed using magnetic bead-based assays, real-time PCR, Western blot, and in-gel mass spectrometry.
• Bovine semen was used to evaluate aptamer interaction with X-spermatozoa.

Main Results:

• Four aptamers (X8, X19, X22, X27) showed preferential binding to X-sorted sperm over Y-sperm in bovine semen.
• Aptamers interacted with a 35 kDa sperm protein, identified as SPACA1, primarily in the acrosome region.
• Competitive Western blot confirmed the differential specificity of aptamers for X-sperm.

Conclusions:

• The study provides novel insights into separating Y-chromosome-bearing sperm from XY populations for producing high-quality cattle.
• Aptamer-based semen sexing is demonstrated as a cost-effective and non-detrimental method for sorting sex-specific spermatozoa.