| Xinjiang province feeds large livestock and plants a great deal of cotton. Vast quantity of by-products of cotton such as cottonseed cake and cottonseed hulls are produced each year, and most of them are used as feedstuff for animals, gossypol poisoning of livestock occur time to time. Chronic gossypol poisoning of sheep and cattle and other livestock represent that blood neutrophils are increased significantly and many tissues and organs are damaged in inflammation. Because interleukin 8 can significantly chemoattract and activate neutrophils and activated neutrophils can lead to serious inflammatory lesion of many tissues and organs, the experiment studied the relationship between significant increase of neutrophils and inflammation of tissues and organs of gossypol poisoning sheep from the expression of interleukin 8.1. Primers of IL-8 of sheep were designed referring to NM001 009 401.1 from NCBI. The complete coding sequence of IL-8 of DuoLang sheep was cloned and sequenced by application of RT-PCR from the extracted total RNA of PBMC. A SNP was found by sequence analysis and it may lead to change of mRNA secondary structure and elevation of protein translation efficiency of IL-8 by analysis of multiple nucleotide sequence alignment and prediction of mRNA secondary structure of IL-8.2. PBMC was stimulated by gossypol concentration of 0,5,10,20,40μmol/L during 12 and 24 hours,then expression of IL-8 was determination by RT-PCR from the extracted total RNA. The experiment shows that expression of IL-8 changes with the gossypol concentration and cell-cultured time and IL-8 may play a leading role in the early inflammation of tissues and organs in gossypol poisoning sheep.3. Prokaryotic expression vector of IL-8 was constructed with pET-28b by DNA recombination upon the construction of IL-8 cloning vector. SDS-PAGE analysis of expressed IL-8 protein laid foundation for further study of IL-8 role in the sheep gossypol poisoning.
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