2021年宁夏新型冠状病毒测序质量分析

Analysis of quality of novel coronavirus sequencing in Ningxia, 2021

目的 分析2021年宁夏新型冠状病毒(简称新冠病毒)测序质量,探索影响因素。方法 2021年10-12月宁夏44份新冠病毒核酸检测阳性样本,以新冠病毒基因组NC_045512.2为参考序列,使用Seqkit、Bowtie 2和Samtools软件统计各样本测序Reads数、基因组覆盖度和平均测序深度,用Ivar软件拼接生成新冠病毒全基因组序列后,使用Quast软件对其质量进行评估。结果 44份样本的ORF1ab基因CT值在14~35,测序Reads数在1.5 M~9.8 M,其中41份样本的基因组覆盖度均在99.0%以上、占93.18%;42份样本拼接后得到完整基因组、占95.45%,基因组分数均在99.00%以上;41份样本的平均测序深度在2 000 X以上、占90.91%。结论 44份样本测序质量较好,病毒载量和病毒序列完整度是影响2021年宁夏新冠病毒测序质量最关键的因素,在基于捕获后进行新冠病毒全基因组测序中,测序Reads数达到1.5 M即可满足基因组拼接需要,继续增加测序Reads数对基因组拼接质量无明显提升。

Objective To analyze the quality of novel coronavirus sequencing in Ningxia, 2021, and explore the influencing factors. Methods Totall 44 positive samples of novel coronavirus PCR detection were selected for genome sequencing.Taking the novel coronavirus genome NC_045512.2 as the reference sequence, the software of Seqkit, Bowtie2 and Samtools were used to count the number of sequencing reads, the coverage and average sequencing depth of genome in each sample.The sequencing quality was evaluated by the software Quast after the genome sequence of novel coronavirus assembled by Ivar software. Results The CT values of ORF1ab genes in 44 samples ranged from 14 to 35, and the number of sequencing reads ranged from 1.5 M to 9.8 M, among which, 41 samples with the genome coverage over 99.0%, accounted for 93.18%,42 samples with complete genome after assembled, accounted for 95.45%, and the Genomic scores were all over 99.00%, 41samples with the average sequencing depth over 2 000 X, accounted for 90.91%. Conclusions The sequencing quality of the 44 samples was good, the viral load and the viral sequence integrity in Ningxia, 2021 are the critical factors affecting the sequencing quality of novel coronavirus. In capture-based whole-genome sequencing of novel coronavirus, the genome splicing meet the needs if the reads number of sequencing reach 1.5 M. There is no significant improvement for the quality of genome splicing, if the reads number of sequencing increases.