Thermostats Gene Amplification Technology Detection Of Salmonella In Animal Feed

Salmonella is a aerobic gram-negative bacteria without Bacillus. It is widely exist in the natural environment and parasitic in the human and animal intestine, infectious gastrointestinal diseases through contaminated food and water, for human and animal health have greater hazards. Animal feed as the Salmonella of non-biological reservoir play an important role in the animal and human disease occurrence and spread of the new bacteria-introduction.In this paper, the three aspects were done: development of LAMP assay to detect Salmonella in animal feed; Study on the role of inner primer in loop-mediated isothermal amplification technology; development of HDA assay to detect Salmonella in animal feed.A highly specific set of primers was synthesized to target the invA gene of Salmonella. Through optimizing the conditions, we have established a LAMP and HDA that can rapidly detect Salmonella . Strains of Salmonella and other pathogens was used to detect the method's specificity . Dilution method was used to assess their sensitivity. In the feed mill near Changsha, collecting 108 farm feed with the new method of detection, at the same time compared to conventional method..The positive detection of Salmonella is the number 20 by the LAMP method, and the positive rate was 18.5% (20/108). The positive detection of Salmonella is the number 19 by the HDA method, and the positive rate was 17.6% (19/108).The conventional method detecte Salmonella is the number 21, and the positive rate was 19.44% (21/108). The two new methods compared with the conventional method, Consistent with the positive rates were 90.48%, 95.24%, in line with the negative rate of 100%.Products were cloned and sequenced evacuation company, the positive results of strains with the cloned DNA sequences in GenBank by the invasion of the same gene sequences. That the establishment of the HAD, LAMP method detection of Salmonella is fast, simple, specific, sensitive features such as a laboratory for the use of the grass-roots level, have broad prospects for promotion.Using PCR-mediated site-directed mutagenesis of the gene of Salmonella invA site-directed mutagenesis to A669→T669, T725→A725, with the original LAMP-specific PCR primers containing single nucleotide sequence differences,Observation of amplification results. PCR results are negative, not the original primer amplification mutation template. LAMP technology can not use the same primers to detect a single nucleotide difference between the same or different plant type plant type of pathogen, and this results associated with the inner primer in the LAMP technology has played a vital role.