18s amplicons covering V4 and V9 hypervariable regions from Anopheles colutzii samples from West Africa (Larval, Unfed adults, Human Bloodfed, Animal Bloodfed) amplified with two PCR clamp approaches to maximize amplification of eukaryotic microbes.

The identification of the eukaryotic microbiota by targeting the 18s rRNA gene is challenging due to simultaneous amplification of the abundant 18s rRNA gene target in the eukaryotic mosquito host. Here, we use defined panels of A. gambiae samples from West Africa to test two experimental PCR clamp approaches to maximize the specific amplification of 18s rRNA gene hypervariable regions from eukaryotic microbes using: i) anneal-inhibiting blocking primers (annealing blockers), and ii) peptide-nucleic acid (PNA) oligonucleotides (PNA blockers).

通过靶向18S核糖体RNA基因（18s rRNA gene）鉴定真核微生物群落颇具挑战，因该方法会同时扩增真核蚊子宿主体内丰度极高的该靶标基因。本研究采用采自西非的标准化冈比亚按蚊（A. gambiae）样本组，对两种实验性PCR钳技术进行测试，以最大化实现对真核微生物18S核糖体RNA基因高变区的特异性扩增，两种技术分别为：i）退火抑制型封闭引物（anneal-inhibiting blocking primers，简称退火封闭引物）；ii）肽核酸（PNA）寡核苷酸（PNA blockers，即PNA封闭剂）。