本研究分别对新老窖泥样本的16S rDNA V4和V3-V4区进行Illumina高通量测序，比较了测序的群落多样性参数、NMDS聚类与优势门/属的相对丰度。V4区的OTUs数目（264~639）显著低于V3-V4区（459~1123）；V4区的Shannon指数在新、老池壁泥中（2.47、5.02）显著低于V3-V4区（3.33、6.46），而在新（4.98vs5.98）、老（5.64vs5.39）池底泥样本中无显著差异（p>0.05）；V4区的Chao1指数在新池底泥（2455vs1358）、新池壁泥（1326vs504）和老池底泥（3475vs1260）中显著高于V3~V4区，在老池壁泥中无显著差异（1915vs1312）（p>0.05）。两个可变区的测序对NMDS聚类规律无影响。热图聚类结果显示，两个可变区的优势门/属的相对丰度很接近，V4区在属水平上更容易区分老窖池的池底泥。与V3-V4区相比，V4区更利于区分池壁泥与池底泥，且可以检测出甲烷菌的含量，更有利于窖泥菌群组成分析。本研究为16S rDNA高通量测序中V3-V4区与V4区的选择提供一定参考。

In this study, Illumina high-throughput sequencing of 16S rDNA V4/V3-V4 variable regions of old and young pit-mud (PM) samples were performed. The diversity indices, NMDS clustering results, and the relative abundance of dominant phyla/genera of two variable regions were compared. The number of OTUs in V4 (264~639) were significantly lower than that of V3-V4 (459~1123). The Shannon indices of V4 in the young (2.47) and old (5.02) Wall-PM (WPM) samples were significantly lower than those of V3-V4 (3.33 and 6.46, respectively). Insignificant difference (p>0.05) was found between the young (4.98vs5.98) and old (5.64vs5.39) bottom-PM (BPM) samples. The Chao1 indices of V4 in the young BPM (2455vs1358), young WPM (1326vs504) and old BPM (3475vs1260) were significantly higher than those of V3-V4, with insignificant difference (p>0.05) in the old WPM samples (1915vs1312). The sequencing of the two variable regions had no effect on the NMDS clustering results. The heat-map clustering results showed that the relative abundance of the dominant phyla/genera for two variable regions were pretty similar, while it was easier for theV4 variable region to distinguish old BPM samples at the genus level. Compare with the V3-V4 region, the V4 variable region was more conducive to distinguishing between WPM and BPM and detecting the content of methanogen, which allows efficient analysis of the PM microbial composition. This study provided a good reference for the selection of V3-V4 and V4 regions during high-throughput sequencing of 16S rDNA.