一株高效降解菜籽饼粕中硫苷的枯草芽孢杆菌 基因组鉴定及功能分析Genome identification and functional analysis of a Bacillus subtilis strain efficiently degrading glucosinolates in rapeseed meal

Abstract

为筛选可高效降解菜籽饼粕中硫苷的细菌，给硫苷降解基因工程菌的构建提供实验数据，以堆放菜籽饼粕的土壤为分离源，通过硫苷选择培养基筛选及硫苷降解率测定，获得一株硫苷降解率最大的菌株C1，采用16S rRNA基因序列进行种属鉴定，通过第3代Illumina NovaSeq PE150测序，获得其基因组完成图并进行功能注释，借助比较基因组学探究C1降解硫苷的机制。结果表明：菌株C1为枯草芽孢杆菌 (Bacillus subtilis)，其硫苷降解率为 (68.98±4.74)%；C1菌株的基因组染色体全长为4 139 381 bp，鸟嘌呤和胞嘧啶含量为43.88%，编码区总长度占全基因组的比例为89.14%；C1菌株能编码大量的功能蛋白，主要涉及氨基酸转运与代谢；比较基因组学分析确认，C1与枯草芽孢杆菌的模式菌株 (NC000964.3) 共线性高，序列中未见大片段易位和倒置的区域，但C1序列中2.7 Mb处出现一个小片段的插入，2.1~2.4 Mb处出现小片段缺失，分别涵盖了2条黑芥子酶的相关基因，推测C1的硫苷降解机制为其可高效分泌黑芥子酶。C1菌株能够高效降解菜籽饼粕中的硫苷，是构建硫苷降解基因工程菌的可选材料。 In order to screen bacteria that can efficiently degrade glucosinolates in rapeseed meal and provide experimental data for the construction of glucosinolate-degrading genetically engineered bacteria, the soil stacked with rapeseed meal was used as the separation source, and a strain C1 with the greatest glucosinolate degradation rate was obtained by glucosinolate selection medium screening and glucosinolate degradation rate determination,and 16S rRNA gene sequence was used for C1 species identification and the third generation Illumina NovaSeq PE150 was used to sequencing the whole genome to obtain C1 genome completion map and functional annotation was carried out. The comparative genomics was used to explore the mechanism for glucosinolate degradation of strain C1. The results showed that strain C1 was Bacillus subtilis, and its glucosinolate degradation rate was (68.98±474)%. The genome chromosome of the strain C1 was 4 139 381 bp in length, and the guanine and cytosine content was 43.88%. The total length of the coding region accounted for 89.14% of the whole genome. Strain C1 could encode a large number of functional proteins, mainly related to amino acid transport and metabolism. Comparative genomics analysis confirmed that C1 was highly colinear with the model strain of Bacillus subtilis (NC000964.3), and there were no large fragment translocation and inversion regions in the sequence. However, a small fragment insertion was found at 2.7 Mb in C1 sequence, and a small fragment deletion appeared at 2.1-2.4 Mb,covering two gens of myrosinase, respectively, and the mechanism of glucosinolate degradation of C1 was presumed to efficiently secrete myrosinase. Strain C1 can efficiently degrade glucosinolates in rapeseed meal and is a potential material for constructing glucosinolate-degrading genetically engineered bacteria.

Keywords

• 菜籽饼粕；硫苷；比较基因组学；枯草芽孢杆菌
• rapeseed meal; glucosinolates; comparative genomics; bacillus subtilis