紫色辣椒花青素降解相关基因的挖掘与分析

doi:10.13304/j.nykjdb.2024.0208

摘要/Abstract

摘要：

为深入研究紫色辣椒花青素降解的分子机制，选用3份紫色辣椒材料（HN191、HN192和HN005），对其花后30 d紫色未熟果实和60 d红色成熟果实进行转录组测序。结果表明，鉴定出7 104个共有的差异表达基因。GO和KEGG富集分析显示，这些共有差异表达基因主要富集在细胞壁、次生代谢、脂质代谢、碳水化合物代谢、氨基酸代谢相关的通路。进一步分析鉴定了3个MBW（MYB-bHLH-WD40）转录因子（包括1个MYB和2个WRKY）、3个过氧化物酶、2个β-葡萄糖苷酶编码基因；通过加权基因共表达网络分析（weighted gene co-expression network analysis，WGCNA）鉴定出1个MADS-box转录因子和1个NAC转录因子。实时荧光定量（quantitative real-time PCR，qRT-PCR）分析表明，这10个基因在红色成熟果实中的表达量均较紫色未熟果实上调，表明这10个候选基因可能与紫色辣椒果实的花青素降解有关。以上研究结果为紫色成熟辣椒的育种提供了潜在基因资源。

关键词: 辣椒, 紫色, 花青素, 降解, 加权基因共表达网络分析

Abstract:

To further study the molecular mechanisms of anthocyanin degradation in purple pepper， the transcriptome data of purple unripe peppers at 30 d after anthesis and red ripe fruits at 60 d after anthesis from 3 varieties （HN191， HN192， HN005） were analyzed. The results showed that 7 104 common differentially expressed genes were identified. GO and KEGG enrichment analysis results inidcated that these genes were predominantly associated with processes such as cell wall， secondary metabolism， lipid metabolism， carbohydrate metabolism and amino acid metabolism. Further analysis identified 3 MBW （MYB-bHLH-WD40） transcription factors （including 1 MYB and 2 WRKY）， 3 peroxidases and 2 β-glucosidase encoding genes. Additionally， a MADS-box transcription factor and a NAC transcription factor were identified through weighted gene co-expression network analysis. The results of qRT-PCR revealed that the expression levels of these genes were higher in red ripe fruits than those in purple unripe fruits， which proposed that these 10 candidate genes might be associated with the degradation of anthocyanins in purple pepper fruits. Above results provided potential candidate genes for the breeding of mature purple peppers.

Key words: chili pepper, purple, anthocyanin, degradation, weighted gene co-expression network analysis

中图分类号:

S641.3

谭华强, 李丽平, 铁曼曼, 杨家勤, 郑晓云, 潘绍坤, 唐有万. 紫色辣椒花青素降解相关基因的挖掘与分析[J]. 中国农业科技导报, 2025, 27(9): 79-91.

Huaqiang TAN, Liping LI, Manman TIE, Jiaqin YANG, Xiaoyun ZHENG, Shaokun PAN, Youwan TANG. Mining and Analysis of Genes Related to Anthocyanin Degradation in Purple Pepper[J]. Journal of Agricultural Science and Technology, 2025, 27(9): 79-91.

图/表 9

表1 候选基因的qRT-PCR分析引物

Table 1 Primers used in qRT-PCR for candidate genes

基因编号 Gene ID	正向引物 Forward primer （5’-3’）	反向引物 Reverse primer（5’-3’）	大小 Size/bp	退火温度 Annealing temperature/℃
T459_29804	TGGAAGAGTTGAACCACCGA	GTGTCCTCGTCACCACTACA	248	59
T459_02736	GGACCAACAACACACCAACA	CTTGGGTCACTGAAGCATCG	249	59
T459_27911	AACGGGTTCTTGGGGCTAAT	GTGCATCGGTAGTAGCTCCT	236	59
T459_07162	CTGCCAGTTGACTTGTTCGT	TGCAAAACGGTCACAGTGAG	249	59
T459_09393	CTGCCAACACTCAAATCCCC	ATTTGCATCGCCACCAGAAG	230	59
T459_33197	TTACCCAACAGTTGACCCGA	CAGCAGCCATTTTCTCCACA	239	59
T459_08317	GTTGCAGCACAGAGAACCAA	ACCAAAGTCCCTGCTATGCT	234	59
T459_08320	TGCTGGTTCTAGATTGCCCA	CTCCTTGCAGACCTGGTTCT	233	59
T459_27108	TGCTTGGAGAGGATTTGGGA	TGTACACTTTGCTCCCCAGG	244	59
T459_12184	ACCCGACTGATGAAGAGCTT	TCGCCTTCCAGTACCCATTT	231	59
T459_30033	CCCTGTCCTGCTCACTGAAG	GTCACGTCCAGCAAGATCCA	250	60

图1 不同品种30与60 d果实发育间的差异表达基因A：差异表达基因直方图； B：差异表达基因的维恩图

Fig. 1 DEGs between fruits of 30 and 60 d from different varietiesA：Histogram of differentially expressed genes； B. Venn plot of differentially expressed genes

图2 共有差异表达基因的富集分析A：GO富集分析；B：KEGG富集分析

Fig. 2 Enrichment analysis of common DEGsA：GO enrichment analysis； B： KEGG enrichment analysis

图3 共有差异表达基因中的MBW转录因子热图注：框中的数值代表FPKM值。

Fig. 3 Heatmap of MBW transcription factors predicted from common DEGsNote：Number in the box indicates FPKM value.

图4 候选基因的相对表达量

Fig. 4 Relative expression of candidate genes

图5 3个辣椒过氧化物酶基因与其他13个Ⅲ类过氧化物酶基因的系统发育树

Fig. 5 Phylogenetic tree of 3 pepper peroxidases and other 13 peroxidase genes of class Ⅲ

图6 加权共表达网络构建A：18个样本的聚类图；B：软阈值筛选；C：基因聚类和模块切割

Fig. 6 Construction of a weighted co-expression networkA：Cluster dendrogram of 18 samples；B：Soft threshold selection；C：Gene clustering and module cutting

图7 WGCNA模块分析A：16个模块所包含的基因数量统计；B：16个模块与花青素含量的相关性热图。*和**分别表示在P<0.05和P<0.01水平显著相关

Fig. 7 Analysis of WGCNA modulesA： Statistics of gene numbers in 16 modules； B： Relationship heatmap of 16 modules with anthocyanin content. * and ** indicate significant correlations at P<0.05 and P<0.01 levels， respectively

图8 WGCNA网络Hub基因分析A： WGCNA得到的蓝色模块degree前50个基因网络图。颜色深浅代表Degree值的大小，颜色越深代表Degree值越高，三角形节点代表转录因子；B：共有差异表达基因PPI网络Degree前50个基因网络图。红色基因名代表与蓝色模块共有的基因，颜色深浅代表Degree值的大小，颜色越深代表degree值越高。三角形节点代表转录因子

Fig. 8 Analysis of hub genes in WGCNA networkA： Network diagram of degree top 50 genes in the blue module obtained by WGCNA. The color shade on the diagram represents the Degree value， with darker colors indicating higher values and triangular node in the diagram represents transcription factor；B： Network diagram of Degree top 50 genes in PPI network of common DEGs. The red gene names indicate genes share with the blue module， the color shade represents the Degree value， with darker colors indicating higher values. Triangular node in this diagram represents transcription factor

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