基于流式细胞术和K-mer分析的黄芪基因组大小估测

目的使用流式细胞术与K-mer分析估测黄芪基因组的大小及复杂程度,为黄芪基因组测序奠定基础。方法以番茄为内标,用流式细胞仪测定经碘化丙啶(PI)染色的番茄细胞核和黄芪细胞核混合样品的PI荧光强度,通过比较黄芪与番茄细胞DNA含量峰值的倍数关系,计算黄芪的基因组大小。采用高通量测序技术进行黄芪基因组调查测序,利用生物信息学方法估计黄芪杂合率、重复序列和GC含量等信息。结果采用流式细胞术测得黄芪基因组大小约为1 426 Mb。通过基因组调查测序,获得了95 Gb黄芪基因组DNA测序数据,K-mer分析表明黄芪基因组约为1 456 Mb,测序深度为39×。K-mer分布曲线有明显的杂合峰,基因组杂合率为2.1%。结论黄芪基因组大小为1.45 Gb左右,杂合率较高。这一结果可为黄芪基因组学研究提供参考。

Estimation of genome sizes of Astragalus membranaceus based on flow cytometry and K-mer analysis

Objective To determinate the genome size and complexity of Astragalus membranaceus by using flow cytometry (FCM) and K-mer analysis, which can lay the foundation for the screening of functional genes of A. membranaceus. Methods Lycopersicon esculentum was served as an internal reference in this study. The mixed sample of A. membranaceus cell nucleus and L. esculentum cell nucleus was stained using propidium iodide (PI). The PI fluorescence intensities of the sample were measured by FCM. The genome size of A. membranaceus was calculated by comparing the multiple relationship between the peak of DNA content in the cells of A. membranaceus and L. esculentum. The genome of A. membranaceus was sequenced by using high-throughput sequencing technologies. The genome size of A. membranaceus was calculated by K-mer analysis. The hybridity percentage, repetitive sequence, and GC of A. membranaceus were estimated by bioinformatics analysis. Results The genome size of A. membranaceus was about 1 426 Mb. For K-mer analysis, more than 95 Gb high quality data from the genome was generated. The average genome size and sequencing coverage depth of A. membranaceus was about 1 456 Mb and 39 times respectively. The genome of A. membranaceus had obvious hybridity peak by K-mer method, and the hybridity percentage as high as 2.1%. Conclusion The genome size of A. membranaceus was about 1.45 Gb and the heterozygosity is high. These data would provide a reference for the genomic research in A. membranaceus.