Determination of the cDNA sequence and mRNA expression of interleukin-1 receptor antagonist in horses.

The researchers determined the structure of a particular horse DNA sequence and its associated RNA sequence, revealing potential differences among horse breeds.

Study Aim and Methods

• The objective of this research was to decipher the complementary DNA (cDNA) sequence for the interleukin-1 receptor antagonist (IL-1ra) in horses, and to compare how the corresponding messenger RNA (mRNA), which is the molecule that interprets DNA instructions to build proteins, expressed this sequence in different horse breeds.
• The researchers utilized samples from the blood of both neonatal (newborn) and adult horses dealing with diverse diseases.
• To identify the precise sequence of the horse DNA, a method known as the polymerase chain reaction was employed. This technique amplified, or increased, the quantity of a specific 220 base pair (bp) segment of the genomic DNA under investigation.
• Surrounding regions of the cDNA sequence – both upstream (5′) and downstream (3′) – were established through a rapid amplification of cDNA ends (RACE) procedure.
• To scrutinize mRNA expression of IL-1ra, a technique known as Northern blot hybridization was utilized.

Study Findings

• The researchers succeeded in establishing the consensus sequence of the cDNA with use of the 5′-RACE procedure, as well as the 220 bp genomic DNA sequence. These sequences are believed to symbolize the sequence secreted for IL-1ra in horses.
• The proposed secreted IL-1ra amino acid sequence, which is built from information in the DNA/ RNA, is composed of 176 residues with an in-frame stop codon. This component of the sequence is considered to represent the signal peptide – a molecule that directs the transportation of proteins – that has been previously reported for human secreted IL-1ra.
• An approximately 1.3 kilobase pair (kb) band, embodying a segment of the 3′ end of the coding region and the 3′ untranslated region, was recognized through application of the 3′ -RACE procedure.
• A 1.6 kb transcript in blood RNA from adult Arabian, Belgian, Thoroughbred, and Standardbred horses was detected using Northern blot hybridization. This suggests that this IL-1ra sequence is common amongst these breeds.
• The results suggest that the DNA encoding for the equine secreted IL-1ra has a 5′ untranslated region of 29 bp, a coding region of 534 bp, and a 3′ untranslated region of approximately 1,080 bp in length.