Transcriptome analysis and non-target-site resistance mechanism of multi-resistant Lolium multiflorum under herbicide stress

Abstract

Lolium multiflorum, one of the most problematic weeds in winter cereal crops, is resistant to herbicidal stress. However, the genome-wide full-length transcripts of L. multiflorum have not been identified, and the mechanism of non-target-site resistance (NTSR) remains unclear. Single-molecule real-time sequencing (SMRT) and RNA-seq technologies were utilised to generate full-length transcripts for mining NTSR related to the detoxification of acetyl-CoA carboxylase (ACCase) and acetolactate synthase (ALS) inhibitor herbicides in multi-resistant L. multiflorum. In total, 65272 and 73799 isoforms from 30562 and 32536 loci were found in the sensitive HNXX01 and resistant HNZMD04 populations, including 44731 and 51792 novel isoforms and 4919 and 5392 novel genes, respectively. In addition, long non-coding RNAs, alternative splicing events, fusion genes, and alternative polyadenylations were found in these two populations. The RNA-seq results showed that 51 and 29 up-regulated differentially expressed genes (DEGs) were co-expressed in the two populations before and after pinoxaden and pyroxsulam treatment, respectively. Quantitative real-time PCR further verified that seven (CYP72A397, Os03g0283200–1, GSTU6–2/3, GSTZ5, UGT1, and SCPL18–1) and six (GSTU6–3/4, GSTZ5, UGT85A24, and SCPL18–1/2) genes were potential metabolising genes for pinoxaden or pyroxsulam, respectively. Furthermore, GSTZ5, GSTU6–3, and SCPL18–1 may be involved in detoxifying pinoxaden and pyroxsulam. These results contribute to the improvement of genome annotation and transcriptomic studies during resistance development and deepen our understanding of multi-resistance mechanisms in weed species under herbicide stress.