园艺学报 ›› 2021, Vol. 48 ›› Issue (5): 860-872.doi: 10.16420/j.issn.0513-353x.2020-0578
收稿日期:
2020-09-08
修回日期:
2021-03-05
出版日期:
2021-05-25
发布日期:
2021-06-07
通讯作者:
罗昌国,王三红
E-mail:376258195@qq.com;wsh3xg@niau.edu.cn
基金资助:
LAN Liming1, LUO Changguo2,*(), WANG Sanhong1,*(
)
Received:
2020-09-08
Revised:
2021-03-05
Online:
2021-05-25
Published:
2021-06-07
Contact:
LUO Changguo,WANG Sanhong
E-mail:376258195@qq.com;wsh3xg@niau.edu.cn
摘要:
为研究湖北海棠[Malus hupehensis(Pamp)Rehd.]响应白粉病菌胁迫过程中相关基因的表达及分子机制,取同一时期爆发白粉病的‘青砧1号’、‘青砧1号’/湖北海棠嫁接苗的接穗部分和湖北海棠等3份材料的叶片,使用BGISEQ-500测序技术对其进行转录组测序分析。结果表明:青砧1号、青砧1号/湖北海棠和湖北海棠对白粉病的抗性依次增加;GO富集显示代谢过程和催化活性相关的GO term在青砧1号、青砧1号/湖北海棠和湖北海棠中被显著富集;KEGG分析显示亚油酸代谢、植物病原体相互作用、磷酸肌醇代谢、油菜素类固醇生物合成和磷脂酰肌醇信号系统等5条代谢途径在‘青砧1号’/湖北海棠和湖北海棠中均显著富集,碳代谢途径只在抗性强的湖北海棠中显著富集。与白粉病抗性相关基因家族表达分析显示,WRKY70、WRKY33、NPR3和WRKY40等转录因子基因在青砧1号/湖北海棠和湖北海棠中较青砧1号显著上调表达。实时荧光定量PCR结果证实了转录组测序结果的准确性。
中图分类号:
兰黎明, 罗昌国, 王三红. 基于转录组测序的湖北海棠抗白粉病机制分析[J]. 园艺学报, 2021, 48(5): 860-872.
LAN Liming, LUO Changguo, WANG Sanhong. Analysis of Resistance Mechanism to Powdery Mildew Based on Transcriptome Sequencing in Malus hupehensis[J]. Acta Horticulturae Sinica, 2021, 48(5): 860-872.
样品名称 Sample name | clean reads数量/Mb Number of clean reads | GC含量/% GC content | Q30/% | 低质量含量/% Low quality content |
---|---|---|---|---|
青砧1号Qingzhen 1 | 46.17(96.40%) | 49.40 | 93.31 | 3.09 |
青砧1号/湖北海棠 Qingzhen 1/Malus hupehensis | 56.53(97.25%) | 47.33 | 93.98 | 2.34 |
湖北海棠 M. hupehensis | 52.78(96.64%) | 48.53 | 93.50 | 2.87 |
表1 转录组测序数据过滤后的质量统计
Table 1 Quality statistics of filtered transcript group sequencing data
样品名称 Sample name | clean reads数量/Mb Number of clean reads | GC含量/% GC content | Q30/% | 低质量含量/% Low quality content |
---|---|---|---|---|
青砧1号Qingzhen 1 | 46.17(96.40%) | 49.40 | 93.31 | 3.09 |
青砧1号/湖北海棠 Qingzhen 1/Malus hupehensis | 56.53(97.25%) | 47.33 | 93.98 | 2.34 |
湖北海棠 M. hupehensis | 52.78(96.64%) | 48.53 | 93.50 | 2.87 |
图 2 青砧1号、青砧1号/湖北海棠和湖北海棠3个材料Unigenes的GO富集分析
Fig. 2 Analysis of gene ontology enrichment of three materials Unigenes of Qingzhen 1,Qingzhen 1/Malus hupehensis and M. hupehensis
图3 青砧1号、青砧1号/湖北海棠和湖北海棠3个材料Unigenes的KEGG富集分析
Fig. 3 Analysis of the KEGG enrichment of three materials Unigenes of Qingzhen 1,Qingzhen 1/Malus hupehensis and M. hupehensis
图4 青砧1号、青砧1号/湖北海棠和湖北海棠3个材料中基因表达量分布
Fig. 4 Distribution of gene expression in three materials of Qingzhen 1,Qingzhen 1/Malus hupehensis and M. hupehensis
基因名称 Gene name | 基因ID Gene ID | Nr-注释 Nr-annotation | 引物序列(5'-3') Primer sequence(5'-3') |
---|---|---|---|
WRKY70 | XP_008388364.1a | WRKY转录因子70[苹果] WRKY transcription factor 70[Malus × domestica] | F:AAACCACCTACATCGGCGAG |
R:ATGAGAATTGACGGTGCGGA | |||
WRKY33 | NP_001315684.1a | WRKY转录因子26[苹果] WRKY transcription factor 26[Malus × domestica] | F:ACAACAACAGCGCCTCTACC |
R:CCCTCCCCAAAAGACAGCAAA | |||
WRKY3 | XP_008352971.1a | WRKY转录因子3[苹果] WRKY transcription factor 3[Malus × domestica] | F:TACACAGGCAACAACAACGC |
R:TCTGATTCCTCGTGCTTGCTT | |||
WRKY22 | XP_008345081.1a | WRKY转录因子22[苹果] WRKY transcription factor 22[Malus × domestica] | F:CTAGTAGTGCTGCCACTGCC |
R:TCCCACCCATAGTTACACCCA | |||
WRKY2 | XP_008344758.1a | WRKY转录因子2[苹果] WRKY transcription factor 2[Malus × domestica] | F:TCTTGGTGCAACGCCTTGAT |
R:TCTCGAGCTCCCAGGAAGAA | |||
NPR3 | ANJ45443.1a | NPR5a [苹果] NPR5a[Malus × domestica] | F:CATCTTCTCCGAAGGACGGG |
R:TCTGTTTGATAAGGAGAGAGCCA | |||
WRKY40 | NP_001315769.1a | WRKY转录因子40[苹果] WRKY transcription factor 40[Malus × domestica] | F:TGCTCAACTCCAAGGCAATG |
R:TTCACTGTCTGCCTTTCGCT | |||
EF1 | MD09G014760b | 内参基因 Reference gene | F:TACTGGAACATCACAGGCTGAC |
R:TGGACCTCTCAATCATGTTGTC |
表2 荧光定量PCR中所用差异表达基因及其引物序列
Table 2 Differentially expressed genes and their primer sequences in qPCR
基因名称 Gene name | 基因ID Gene ID | Nr-注释 Nr-annotation | 引物序列(5'-3') Primer sequence(5'-3') |
---|---|---|---|
WRKY70 | XP_008388364.1a | WRKY转录因子70[苹果] WRKY transcription factor 70[Malus × domestica] | F:AAACCACCTACATCGGCGAG |
R:ATGAGAATTGACGGTGCGGA | |||
WRKY33 | NP_001315684.1a | WRKY转录因子26[苹果] WRKY transcription factor 26[Malus × domestica] | F:ACAACAACAGCGCCTCTACC |
R:CCCTCCCCAAAAGACAGCAAA | |||
WRKY3 | XP_008352971.1a | WRKY转录因子3[苹果] WRKY transcription factor 3[Malus × domestica] | F:TACACAGGCAACAACAACGC |
R:TCTGATTCCTCGTGCTTGCTT | |||
WRKY22 | XP_008345081.1a | WRKY转录因子22[苹果] WRKY transcription factor 22[Malus × domestica] | F:CTAGTAGTGCTGCCACTGCC |
R:TCCCACCCATAGTTACACCCA | |||
WRKY2 | XP_008344758.1a | WRKY转录因子2[苹果] WRKY transcription factor 2[Malus × domestica] | F:TCTTGGTGCAACGCCTTGAT |
R:TCTCGAGCTCCCAGGAAGAA | |||
NPR3 | ANJ45443.1a | NPR5a [苹果] NPR5a[Malus × domestica] | F:CATCTTCTCCGAAGGACGGG |
R:TCTGTTTGATAAGGAGAGAGCCA | |||
WRKY40 | NP_001315769.1a | WRKY转录因子40[苹果] WRKY transcription factor 40[Malus × domestica] | F:TGCTCAACTCCAAGGCAATG |
R:TTCACTGTCTGCCTTTCGCT | |||
EF1 | MD09G014760b | 内参基因 Reference gene | F:TACTGGAACATCACAGGCTGAC |
R:TGGACCTCTCAATCATGTTGTC |
图5 青砧1号、青砧1号/湖北海棠和湖北海棠3个材料中差异基因表达量分析与验证
Fig. 5 Analysis and verification of differentially expressed genes in three materials of Qingzhen 1,Qingzhen 1/Malus hupehensis and M. hupehensis
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