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为了建立一种特异、敏感、简便的疟疾诊断方法，设计并合成两对针对恶性疟原虫（Ｐ．ｆ．）小亚单位核糖体核糖核酸（ＳＳＵｒＲＮＡ）基因特异的引物，采用套式聚合酶链式反应（ＰＣＲ）技术，扩增恶性疟原虫ＳＳＵｒＲＮＡ基因特定片段。利用该系统诊断云南及海南疟疾患者血样，并随机选取扩增产物进行克隆测序鉴定。结果显示，恶性疟原虫模板均扩增出长度为２０５ｂｐ的特定基因片段，经Ｔ－载体克隆测序与恶性疟原虫ＳＳＵｒＲＮＡ特定基因片段序列完全符合。该系统特异性强，除恶性疟原虫外，间日疟原虫（Ｐ．ｖ．）、三日疟原虫（Ｐ．ｍ．）、卵形疟原虫（Ｐ．ｏ．）、正常人血ＤＮＡ样本及空白对照均无此扩增带出现。该系统检测原虫的敏感度为１．５个原虫／μｌ，大大高于常规镜检。６１份Ｐ．ｆ．镜检阳性标本在进行ＰＣＲ检测时亦均呈阳性，同时联合应用套式ＰＣＲ扩增间日疟原虫ＳＳＵｒＲＮＡ特定基因片段的检测系统，发现６１例恶性疟病人中２３例是Ｐ．ｆ．和Ｐ．ｖ．混合感染。该检测系统具有特异、敏感、稳定、简便的特点，对疟疾的诊断、混合感染的判定具有一定的临床应用价值
In order to establish a specific, sensitive and simple diagnostic method for malaria, two pairs of primers specific to the P.f. small subunit ribosomal RNA (SSUrRNA) gene were designed and synthesized, and nested polymerase chain Type reaction (PCR) technique to amplify specific fragments of the SSUrRNA gene of Plasmodium falciparum. The system was used to diagnose malaria patients in Yunnan and Hainan and randomly selected amplification products for cloning and sequencing. The results showed that the specific gene fragment of 205bp in length was amplified from P. falciparum template, and the sequence of T-vector cloned was in good agreement with the sequence of SSUrRNA specific gene of P. falciparum. This system is highly specific, in addition to Plasmodium falciparum, P. v. P., P. malaria (P.m.), P. o., normal human blood DNA No sample and blank control amplification bands appear. The system detects parasite sensitivity of 1.5 protozoa / μl, much higher than the conventional microscopy. 61 p. f. The positive samples of microscopic examination were all positive when PCR test was performed. In addition, the detection system of specific gene fragment of SSUrRNA of Plasmodium vivax was amplified by nested PCR, and 23 cases of 61 malaria cases were found. f. And P. v. Mixed infection. The detection system has the characteristics of being specific, sensitive, stable and simple, and has certain clinical application value for the diagnosis of malaria and the determination of mixed infection