Analysis Of Numb Taste And Aroma Quality Of Different Zanthoxylum Bungeanum Maxim.Germplasm Resources And Screening Of Key Genes For Sanshool Synthesis

Zanthoxylum bungeanum Maxim.originated in China,widely distributed in China,a wide variety,is an important woody spices and medicinal plants.The appearance of Prickly ash from different producing areas is similar,but the economic application value and quality are quite different.In this study,SSR molecular markers were used to distinguish different prickly ash germplasm resources.Through the analysis of numb and aroma quality and sanshool gene screening of representative Prickly ash germplasm resources,To select highquality Prickly ash varieties and accelerate the healthy development of the industry.It provides a theoretical basis for Prickly ash breeding and sanshool functional gene research.The main results of this study are as follows :1.Identification of different Prickly ash germplasm resources by SSR molecular markersA total of 76 loci were amplified from 38 different Prickly ash germplasms by 16 pairs of primers.The genetic distance of 38 Prickly ash samples was calculated by NTSYSpc Version 2.1 software to be 0 — 1.6675.In the cluster analysis,38 samples were divided into two categories when the genetic similarity coefficient was 0.54,and 38 samples were divided into four categories when the genetic similarity coefficient was 0.67.2.Content and composition analysis of volatile oil from different Prickly ash germplasm resources(1)The volatile oil content of 15 representative Prickly ash samples was determined.The volatile oil content of different Prickly ash germplasm resources was quite different.The volatile oil content of cultivated varieties ranged from 3.8 m L/100 g to 7.77 m L/100 g,with an average content of 5.9 m L/100 g.The volatile oil content of Shi Zi Tou and Wu Ci Mei Hua Jiao was relatively high,reaching 7.77 m L/100 g and 6.95 m L/100 g.The volatile oil content of Zanthoxylum simulans Hance.was only 1.04 m L/100 g.The volatile oil content of wild pepper was significantly lower than that of cultivated pepper.(2)A total of 123 volatile oil components were detected in 15 Prickly ash samples,which were mainly terpenes,alcohols and esters,with a total content of more than 97 %.Except for the highest content of Fu You Jiao esters,the other Prickly ash samples had the highest content of terpenes.Through cluster analysis,it was found that the volatile oil components of different varieties of Prickly ash were significantly different.3.Analysis of sanshool content in different Prickly ash germplasm resourcesThe content of sanshool in 15 representative samples of Prickly ash was determined.There was significant difference in sanshool in different varieties of Prickly ash The sanshool content of cultivated Prickly ash ranged from 5.65 mg/g — 22.87 mg/g,the average content was 16.23 mg/g,and the content of Huang Gai and Wu Ci Mei Hua Jiao was relatively high,reaching 22.87 mg/g and 21.48 mg/g respectively,The content of sanshool in Zanthoxylum simulans Hance was only 4.34 mg/g.The content of numb taste in wild pepper was significantly lower than that in cultivated pepper.4.Gene screening of sanshool(1)The difference of sanshool content between Huang Gai and Shao Ci Da Hong Pao was large,and the difference of volatile oil content was small.Only sanshool gene was analyzed here.,The gene expression levels of Huanggai and Shaoci Dahongpao were analyzed by transcriptomics.The results showed that there were 30,269 differentially expressed genes in Huang Gai and Shao Ci Da Hong Pao(the difference multiple was greater than 1.1 and FDR < 0.01 was used as the screening condition),including 15,136 up-regulated genes and 15,133 down-regulated genes.Finally,five differentially expressed genes related to sanshool production were screened.(2)The differentially expressed genes(EVM0048594,EVM0050045,EVM0023274,EVM0024855,EVM004405)were screened as the target genes,and the candidate genes were verified by real-time PCR.The results showed that the quantitative expression of EVM0050045 and EVM0024855 was consistent with the content of sanshool.(3)To further validate EVM0050045 and EVM0024855 genes,transfer them into tobacco and detect the content of sanshool,the results showed that the content of sanshool in transgenic tobacco plants with EVM0050045 gene and EVM0024855 gene was significantly higher than that in control plants.Therefore,it can be determined that EVM0050045 gene and EVM0024855 gene will promote the production of sanshool.