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作物学报 ›› 2024, Vol. 50 ›› Issue (6): 1503-1513.doi: 10.3724/SP.J.1006.2024.34172

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

不同马铃薯淀粉含量差异的转录组学解析

赵娜1,2(), 刘宇曦1,2, 张朝澍1,2,*(), 石瑛1,2,*()   

  1. 1东北农业大学农学院, 黑龙江哈尔滨 150030
    2寒地粮食作物种质创新与生理生态教育部重点实验室, 黑龙江哈尔滨 150030
  • 收稿日期:2023-10-20 接受日期:2024-01-31 出版日期:2024-06-12 网络出版日期:2024-02-21
  • 通讯作者: * 石瑛, E-mail: shiying01@163.com; 张朝澍, E-mail: zhangchaoshu@126.com
  • 作者简介:E-mail: zhaona992024@163.com
  • 基金资助:
    黑龙江省“揭榜挂帅”科技攻关项目(2022ZXJ06B02);黑龙江省自然科学基金项目(LH2021C027)

Transcriptomic analysis of differences in the starch content of different potatoes

ZHAO Na1,2(), LIU Yu-Xi1,2, ZHANG Chao-Shu1,2,*(), SHI Ying1,2,*()   

  1. 1College of Agronomy, Northeast Agricultural University, Harbin 150030, Heilongjiang, China
    2Key Laboratory of Germplasm Enhancement, Physiology and Ecology of Food Crops in Cold Region, Ministry of Education, Harbin 150030, Heilongjiang, China
  • Received:2023-10-20 Accepted:2024-01-31 Published:2024-06-12 Published online:2024-02-21
  • Contact: * E-mail: shiying01@163.com;E-mail: zhangchaoshu@126.com
  • Supported by:
    “Open Bidding for Selecting the Best Candidates” Scientific and Technological Research Project of Heilongjiang Province(2022ZXJ06B02);Natural Science Foundation of Heilongjiang Province(LH2021C027)

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摘要：

淀粉是马铃薯重要的品质性状之一, 广泛应用于食品、医药、石油化工等行业, 市场上对高淀粉马铃薯品种的需求逐年提高。为探究马铃薯块茎淀粉积累特性及调控的关键基因, 本研究通过RNA测序分析, 分别对高、低淀粉含量马铃薯品种大西洋(DXY)、定薯1号(DS1)的匍匐茎(a)、块茎膨大前期(b)、块茎膨大中期(c)、块茎膨大后期(d)及成熟期(e)进行转录组谱分析, 5个时期共获得9494个差异表达基因(DEGs), 结合功能注释发现差异基因主要富集在结合绑定和催化活性等分子功能中, 代谢通路分析发现差异基因主要富集在碳水化合物途径中, 其中有137个DEGs与淀粉和蔗糖代谢相关。筛选获得9个调控淀粉合成关键基因, 其中蔗糖合酶基因PGSC0003DMG400013547在大西洋整个生育期表达量较高, 且在各时期表达量均高于定薯1号; 果糖激酶基因PGSC0003DMG400026916在a、b时期大西洋的表达量显著高于定薯1号; 定薯1号中淀粉酶基因PGSC0003DMG400009891、PGSC0003DMG400001549和葡聚糖内切-1,3-β-葡萄糖苷酶基因PGSC0003DMG400024642、PGSC0003DMG400003181分别在c和e时期表达量显著高于高淀粉品种大西洋。上述基因可能是参与淀粉的合成与积累的关键调控基因。本研究为不同类型马铃薯品种块茎淀粉代谢调控机制研究提供参考。

关键词: 马铃薯, 发育时期, 淀粉合成积累, 转录组测序

Abstract:

Starch is one of the most important quality characteristics of potatoes, which is widely used in the food, medical, petrochemical and other industries. The demand for starchy potato varieties in the market is increasing year by year. To explore the characteristics and key genes of starch accumulation and regulation of potato tuber, in this study, transcriptome profiling of creeping stems (a), pre-tuber expansion (b), mid-tuber expansion (c), late tuber expansion (d), and mature stage (e) of high and low starch potato cultivars DXY and DS1 were conducted by RNA sequencing analysis, and a total of 9494 differentially expressed genes (DEGs) were identified at five stages. Binding function annotation revealed that the differential genes were mainly enriched in molecular functions such as binding and catalytic activity. Metabolic pathway analysis revealed that the differential genes were mainly enriched in carbohydrate-related metabolic pathways, and 137 DEGs were associated with starch and sucrose metabolism. Nine key genes regulating starch synthesis were examined, and the relative expression level of the sucrose synthase gene PGSC0003DMG400013547 was higher during growth period of DXY than that at any stage. The relative expression level of the fructokinase gene PGSC0003DMG400026916 was significantly higher in DXY than in DS1 during the a and b periods. The relative expression level of amylase gene PGSC0003DMG400009891, PGSC0003DMG400001549, and glucan endo-1,3-β-glucosidase gene PGSC0003DMG400024642, PGSC0003DMG400003181 in DS1 was significantly higher than that in DXY with starch-rich cultivars during the c and e periods. The described gene might be a key regulatory gene for starch synthesis and accumulation. This study provides a clue for the investigation of the regulatory mechanism of tuber starch metabolism in different potato cultivars.

Key words: potato, developmental period, accumulation of starch synthesis, transcriptome sequencing

表1

引物序列"

基因编号Gene ID 正向引物Forward primer (5'-3') 反向引物Reverse primer (5'-3')
PGSC0003DMG400026916 TGATCAAGGTCAGCGATGTG GTCTTCACGTGGAATCCTCC
PGSC0003DMG400016481 ACAGTTCTGCGTTTCAGAGC AGCAGCATGGTAAAGCTCCAA
PGSC0003DMG400013547 ACAAGGGGTGCTTTCGTTCA GTAGCCTTGTCGCCCTGATT
PGSC0003DMG400003181 CGCTGAGAGGAATTTCGGGT TTTGGTTGCCCTTTGTTGCC
PGSC0003DMG400024642 TACAAACGGACCTTGCTTTA AGTAGCAGGATAAACACAGC
PGSC0003DMG400009891 GATCATATTGCAGGCATTCG CAAGCAGTTGTGAAACCAG
EF1a GATGTTGTGCCAAAGGATGT AACTTGGTCAATGCGAGA

图1

定薯1号、大西洋块茎不同发育时期图 Da~De: 定薯1号定植后25 d、40 d、61 d、82 d和103 d; Xa~Xe: 大西洋定植后25 d、40 d、61 d、82 d和103 d。"

图2

定薯1号vs大西洋不同时期淀粉含量和淀粉含量比率的变化 A: 淀粉含量柱状图; B: 淀粉含量比率折线图。Da~De: 定薯1号定植后25 d、40 d、61 d、82 d和103 d; Xa~Xe: 大西洋定植后25 d、40 d、61 d、82 d和103 d。星号表示通过单因素方差分析与定薯1号相比具有显著性(** P < 0.01, *** P < 0.001), ns表示无显著差异(P > 0.05)。"

表2

测序数据质量统计"

样品名称
Sample name		 有效数据
Clean reads		 基因组比对
Mapped reads		 基因组比对率
Mapped reads (%)		 GC含量
GC content (%)		 ≥Q20 (%) ≥Q30 (%)
Da-1 41,710,042 35,971,385 86.24 42.61 97.77 93.98
Da-2 43,243,492 37,736,387 87.26 42.56 97.67 93.48
Da-3 38,555,946 34,064,046 88.35 42.86 97.96 94.21
Db-1 40,147,012 34,518,026 85.98 42.47 97.95 94.06
Db-2 41,452,450 35,287,769 85.13 43.29 97.64 93.87
Db-3 41,485,436 35,915,281 86.57 42.84 98.07 94.60
Dc-1 38,488,084 30,296,884 78.72 41.65 97.88 93.99
Dc-2 38,988,180 30,707,745 78.76 41.46 97.99 94.24
Dc-3 40,364,158 34,737,460 86.06 42.59 98.10 94.55
Dd-1 39,833,928 31,152,181 78.21 41.68 97.96 94.32
Dd-2 41,331,412 32,215,855 77.95 41.58 97.97 94.34
Dd-3 41,865,794 33,566,589 80.18 41.71 97.85 93.99
De-1 41,161,928 35,155,400 85.41 44.47 97.31 93.57
De-2 38,363,322 29,762,706 77.58 42.16 97.46 93.60
De-3 42,150,428 32,190,017 76.37 42.13 97.71 93.98
Xa-1 40,619,164 35,357,453 87.05 42.54 98.28 94.87
Xa-2 40,601,792 35,382,889 87.15 42.68 97.95 94.07
Xa-3 41,200,382 35,910,567 87.16 42.65 97.67 93.37
Xb-1 44,599,490 38,997,447 87.44 42.85 98.26 94.79
Xb-2 41,602,324 35,537,658 85.42 42.59 98.12 94.52
Xb-3 42,202,702 35,250,119 83.53 43.60 97.99 94.49
Xc-1 40,274,444 31,599,634 78.46 42.08 98.18 94.72
Xc-2 39,511,076 32,460,237 82.15 42.20 97.88 94.03
Xc-3 40,493,952 31,485,795 77.75 41.53 98.09 94.57
Xd-1 40,558,872 32,781,195 80.82 41.94 98.16 94.59
Xd-2 40,557,808 32,991,680 81.34 41.88 98.31 94.91
Xd-3 40,031,264 30,777,525 76.88 41.10 98.02 94.18
Xe-1 40,503,764 33,015,497 81.51 42.09 98.24 94.84
Xe-2 41,759,668 34,419,851 82.42 42.56 98.13 94.62
Xe-3 39,888,416 31,042,204 77.82 41.33 98.10 94.51

图3

定薯1号vs大西洋不同时期差异基因比较 A: 差异基因火山图; B: 差异基因数目统计图; C: 差异基因韦恩图。Da~De: 定薯1号定植后25 d、40 d、61 d、82 d和103 d; Xa~Xe: 大西洋定植后25 d、40 d、61 d、82 d和103 d。"

图4

定薯1号vs大西洋差异基因富集分析 A: 差异表达基因的GO分类; B: 前20条最显著富集的KEGG途径。"

图5

定薯1号vs大西洋不同时期差异基因KEGG富集分析 Da~De: 定薯1号定植后25 d、40 d、61 d、82 d和103 d; Xa~Xe: 大西洋定植后25 d、40 d、61 d、82 d和103 d。"

图6

不同时期淀粉和蔗糖代谢通路相关差异基因相对表达量热图 用DESeq2 log2 (定薯1号 vs 大西洋)表示各基因的相对表达量, 浅蓝色表示上调, 深蓝色表示下调。Da~De: 定薯1号定植后25 d、40 d、61 d、82 d和103 d; Xa~Xe: 大西洋定植后25 d、40 d、61 d、82 d和103 d。"

图7

两品种不同时期差异基因表达量热图分析 用log2 (FPKM)表示基因表达量, 红色表示高表达, 蓝色表示低表达。Da~De: 定薯1号定植后25 d、40 d、61 d、82 d和103 d; Xa~Xe: 大西洋定植后25 d、40 d、61 d、82 d和103 d。"

图8

差异基因qRT-PCR验证 A: qRT-PCR与RNA-seq结果比较, B: qRT-PCR与RNA-seq相关性分析。Da~De: 定薯1号定植后25 d、40 d、61 d、82 d和103 d; Xa~Xe: 大西洋定植后25 d、40 d、61 d、82 d和103 d。"

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