猪源2型链球菌福建株cps2j基因PCR检测及序列分析

PCR Detection and Sequence Analysis of cps2j Gene for Fujian Strain Streptococcus Suis Serotype 2

设计并合成一对扩增2型猪链球菌cps基因的特异性引物,以猪2型链球菌福建株(SS2PFJ07)DNA为模板,筛选最佳反应条件,建立检测cps2j基因的PCR方法,并对PCR扩增产物进行序列测定和同源性比较分析。结果如下:应用该方法对猪2型链球菌福建株和标准阳性株进行扩增,均获得与预期大小一致的675bp特异性目的片段,而对8株非2型猪链球菌的扩增结果均呈阴性;敏感性测定最低可检出100cfu细菌量或50pg细菌DNA;SS2PFJ07cps2j基因部分核苷酸及其推导的氨基酸序列与猪链球菌2型不同菌株的同源性分别达98.7%～100%和97.8%～100%。上述结果表明建立并优化的猪2型链球菌cps2j基因的PCR检测方法敏感性好、特异性高,能用于2型猪链球菌的分子流行病学调查和快速检测;序列分析表明猪2型链球菌福建株与国内外8株标准株之间同源性高、亲缘关系密切。

To synthesize a pair of specific primers,the PCR method was established to detect the Streptococcus suis serotypes 2（SS2） cps2j partial gene by means of the SS2 Fujian（PFJ07） strain DNA as a template,and the product（cps2j partial gene） of PCR was sequenced and blasted. The results were as follows：The 675bp target fragment was amplified by the PCR from the SS2 PFJ07 strain and the standard positive SS2. but other 8 strains Streotococcus suis but not SS2 were not amplified. The sensitive test indicated the PCR less can detect 100cfu of bacterium quantity or 50pg of genic DNA. The sequence of PCR product achieved to 98%-100% nucleotide homology with that of SS2 other strains,amino acid sequence achieved to 91.6%-93.3%. The results revealed the PCR developed was sensitive and specific and can be applied to the SS2 diagnosis and epidemiology investigation. Sequence analysis indicated homology between the SS2 PFJ07 strain and 8 SS2 at home and abroad was high and genetic relationship was close.