园艺学报 ›› 2021, Vol. 48 ›› Issue (5): 849-859.doi: 10.16420/j.issn.0513-353x.2020-0895
• 研究论文 • 下一篇
姚富文1,*, 王枚阁1,*, 宋春晖1, 宋尚伟1, 焦健1, 王苗苗1, 王昆2, 白团辉1,**(), 郑先波1,**(
)
收稿日期:
2020-12-29
修回日期:
2021-04-08
出版日期:
2021-05-25
发布日期:
2021-06-07
通讯作者:
白团辉,郑先波
E-mail:tuanhuibai88@163.com;xianboz@163.com
作者简介:
*共同第一作者
基金资助:
YAO Fuwen1,*, WANG Meige1,*, SONG Chunhui1, SONG Shangwei1, JIAO Jian1, WANG Miaomiao1, WANG Kun2, BAI Tuanhui1,**(), ZHENG Xianbo1,**(
)
Received:
2020-12-29
Revised:
2021-04-08
Online:
2021-05-25
Published:
2021-06-07
Contact:
BAI Tuanhui,ZHENG Xianbo
E-mail:tuanhuibai88@163.com;xianboz@163.com
摘要:
为了探究热激蛋白(HSP90)基因家族成员在苹果(Malus × domesticaBorkh.)耐热中的作用,利用生物信息学方法对苹果HSP90基因家族成员进行鉴定,并分析其在高温胁迫下和不同组织中的表达规律。结果表明,苹果HSP90基因家族有11个成员,分布于10条染色体上,其氨基酸序列大小介于104 ~ 818 aa,蛋白质分子质量介于11.81 ~ 93.58 kD。转录组分析显示,11个MdHSP90家族成员中,有4个基因,MdHSP90-1、MdHSP90-3、MdHSP90-5和MdHSP90-11在高温胁迫下显著上调;亚细胞定位显示这4个基因中前三者定位在细胞膜与细胞核上,而后者定位在细胞核上;进一步通过qRT-PCR分析发现这4个基因在两种苹果砧木平邑甜茶(Malus hupehensis)和变叶海棠(Malus toringoides)中的表达趋势基本一致,其表达量在高温胁迫早期大幅升高,在中后期有所下降;不同苹果组织中均能检测到这4个基因的表达,高温处理可不同程度地激活这些基因的表达,表明它们可能在苹果耐热胁迫中具有重要作用。
中图分类号:
姚富文, 王枚阁, 宋春晖, 宋尚伟, 焦健, 王苗苗, 王昆, 白团辉, 郑先波. 苹果HSP90家族基因鉴定及高温胁迫下的表达分析[J]. 园艺学报, 2021, 48(5): 849-859.
YAO Fuwen, WANG Meige, SONG Chunhui, SONG Shangwei, JIAO Jian, WANG Miaomiao, WANG Kun, BAI Tuanhui, ZHENG Xianbo. Identification and Expression Analysis of HSP90 Gene Family Under High Temperature Stress in Apple[J]. Acta Horticulturae Sinica, 2021, 48(5): 849-859.
用途Useage | 基因 Gene | 引物(5′-3′)Primer |
---|---|---|
实时荧光定量qRT-PCR | MdHSP90-1 | F:CGTCGATTCGGATGATTTGC;R:TGGCTCTGTTCTGGCTGTCC |
MdHSP90-3 | F:GGAAATCAGCGACGACGAAG;R:CAAGGTGCTCCTCCCAGTCA | |
MdHSP90-5 | F:ATGTTGCTTCTGACAGCGTGAC;R:CCATATTCCTGCTTAATCTCCTTTT | |
MdHSP90-11 | F:CGTATTGCTCCGTTGCTTAG;R:AGTTTCTCCAGGAATGGTGTATT | |
Actin | F:GGATTTGCTGGTGATGATGCT;R:AGTTGCTCACTATGCCGTGC | |
亚细胞定位 Subcellular location | GFPHSP90-1 | F:GTGGATCCAAAGAATTCATGGCCGACGTTCAGATGCAC |
R:CTCCTTTACCCATGAATTCTCAAAATTATATTAACATTATCGAG | ||
GFPHSP90-3 | F:GTGGATCCAAAGAATTCCCGATGTCAGCATGATTGG | |
R:CTCCTTTACCCATGAATTCAAAAACCCGAAACATCCACAAACTC | ||
GFPHSP90-9 | F:GTGGATCCAAAGAATTCATGCACAGGCTCCCAC | |
R:CTCCTTTACCCATGAATTCTTTCTGGCTGCC | ||
GFPHSP90-11 | F:GTGGATCCAAAGAATTCATGCACAGGCTCCCACGACGCTCCG | |
R:CTCCTTTACCCATGAATTCTTTCTGACTGCCG |
表1 qRT-PCR和亚细胞定位引物序列
Table 1 Primer sequences of qRT-PCR and subcellular location
用途Useage | 基因 Gene | 引物(5′-3′)Primer |
---|---|---|
实时荧光定量qRT-PCR | MdHSP90-1 | F:CGTCGATTCGGATGATTTGC;R:TGGCTCTGTTCTGGCTGTCC |
MdHSP90-3 | F:GGAAATCAGCGACGACGAAG;R:CAAGGTGCTCCTCCCAGTCA | |
MdHSP90-5 | F:ATGTTGCTTCTGACAGCGTGAC;R:CCATATTCCTGCTTAATCTCCTTTT | |
MdHSP90-11 | F:CGTATTGCTCCGTTGCTTAG;R:AGTTTCTCCAGGAATGGTGTATT | |
Actin | F:GGATTTGCTGGTGATGATGCT;R:AGTTGCTCACTATGCCGTGC | |
亚细胞定位 Subcellular location | GFPHSP90-1 | F:GTGGATCCAAAGAATTCATGGCCGACGTTCAGATGCAC |
R:CTCCTTTACCCATGAATTCTCAAAATTATATTAACATTATCGAG | ||
GFPHSP90-3 | F:GTGGATCCAAAGAATTCCCGATGTCAGCATGATTGG | |
R:CTCCTTTACCCATGAATTCAAAAACCCGAAACATCCACAAACTC | ||
GFPHSP90-9 | F:GTGGATCCAAAGAATTCATGCACAGGCTCCCAC | |
R:CTCCTTTACCCATGAATTCTTTCTGGCTGCC | ||
GFPHSP90-11 | F:GTGGATCCAAAGAATTCATGCACAGGCTCCCACGACGCTCCG | |
R:CTCCTTTACCCATGAATTCTTTCTGACTGCCG |
基因名称 Gene name | 序列ID Sequence ID | 染色体 Chr | 基因组位置 Genomic location | 开放阅读框/bp ORF | 氨基酸/aa Amino acid | 蛋白质分子质量/kD mw | 等电点 pI |
---|---|---|---|---|---|---|---|
MdHSP90-1 | MD01G1208700 | 1 | 30 315 406 ~ 30 318 680 | 2 112 | 703 | 80.77 | 5.00 |
MdHSP90-2 | MD03G1036600 | 3 | 2 907 372 ~ 2 910 570 | 2 100 | 699 | 80.21 | 4.95 |
MdHSP90-3 | MD07G1279200 | 7 | 34 367 607 ~ 34 369 506 | 1 749 | 582 | 67.39 | 5.06 |
MdHSP90-4 | MD08G1011200 | 8 | 864 457 ~ 869 106 | 2 547 | 818 | 93.59 | 4.85 |
MdHSP90-5 | MD09G1122200 | 9 | 9 421 880 ~ 9 427 733 | 2 430 | 809 | 91.89 | 5.29 |
MdHSP90-6 | MD11G1037400 | 11 | 3 204 284 ~ 3 207 461 | 2 100 | 699 | 80.04 | 4.95 |
MdHSP90-7 | MD11G1152300 | 11 | 14 614 376 ~ 14 614 690 | 315 | 104 | 11.81 | 4.16 |
MdHSP90-8 | MD13G1055300 | 13 | 3 867 643 ~ 3 871 210 | 1 143 | 380 | 43.56 | 4.89 |
MdHSP90-9 | MD14G1110600 | 14 | 17 566 378 ~ 17 567 467 | 930 | 309 | 36.35 | 9.40 |
MdHSP90-10 | MD16G1054100 | 16 | 3 825 148 ~ 3 830 828 | 2 385 | 794 | 89.67 | 5.02 |
MdHSP90-11 | MD17G1113200 | 17 | 9 690 370 ~ 9 696 630 | 2 421 | 806 | 91.69 | 5.34 |
表2 苹果HSP90基因家族成员的理化性质
Table 2 Physical and chemical properties of HSP90 gene family in apple
基因名称 Gene name | 序列ID Sequence ID | 染色体 Chr | 基因组位置 Genomic location | 开放阅读框/bp ORF | 氨基酸/aa Amino acid | 蛋白质分子质量/kD mw | 等电点 pI |
---|---|---|---|---|---|---|---|
MdHSP90-1 | MD01G1208700 | 1 | 30 315 406 ~ 30 318 680 | 2 112 | 703 | 80.77 | 5.00 |
MdHSP90-2 | MD03G1036600 | 3 | 2 907 372 ~ 2 910 570 | 2 100 | 699 | 80.21 | 4.95 |
MdHSP90-3 | MD07G1279200 | 7 | 34 367 607 ~ 34 369 506 | 1 749 | 582 | 67.39 | 5.06 |
MdHSP90-4 | MD08G1011200 | 8 | 864 457 ~ 869 106 | 2 547 | 818 | 93.59 | 4.85 |
MdHSP90-5 | MD09G1122200 | 9 | 9 421 880 ~ 9 427 733 | 2 430 | 809 | 91.89 | 5.29 |
MdHSP90-6 | MD11G1037400 | 11 | 3 204 284 ~ 3 207 461 | 2 100 | 699 | 80.04 | 4.95 |
MdHSP90-7 | MD11G1152300 | 11 | 14 614 376 ~ 14 614 690 | 315 | 104 | 11.81 | 4.16 |
MdHSP90-8 | MD13G1055300 | 13 | 3 867 643 ~ 3 871 210 | 1 143 | 380 | 43.56 | 4.89 |
MdHSP90-9 | MD14G1110600 | 14 | 17 566 378 ~ 17 567 467 | 930 | 309 | 36.35 | 9.40 |
MdHSP90-10 | MD16G1054100 | 16 | 3 825 148 ~ 3 830 828 | 2 385 | 794 | 89.67 | 5.02 |
MdHSP90-11 | MD17G1113200 | 17 | 9 690 370 ~ 9 696 630 | 2 421 | 806 | 91.69 | 5.34 |
图4 苹果HSP90基因在高温胁迫下的表达分析 PCK:平邑甜茶对照;PH:平邑甜茶高温;BCK:变叶海棠对照;BH:变叶海棠高温。 不同小写字母表示同一种砧木在不同胁迫时间表达量存在显著差异(P < 0.05)。
Fig. 4 Expression analysis of HSP90 genes in apple under high temperature stress PCK:Malus hupehensis(control);PH:Malus hupehensis(high temperature);BCK:Malus toringoides(Control);BH:Malus toringoides(high temperature). Different lowercase letters in each bar indicate significant differences at different stress times in the same rootstock(P < 0.05).
图 5 苹果HSP90基因的组织表达分析 不同小写字母表示同一种砧木不同组织之间表达量存在显著差异(P < 0.05)。
Fig. 5 Tissue expression analysis of HSP90 genes in apple In the same apple rootstock,different lowercase letters indicated significant differences in the expression levels among different tissues(P < 0.05).
图 6 MdHSP90-1、MdHSP90-3、MdHSP90-5和MdHSP90-11蛋白在烟草细胞中的亚细胞定位
Fig. 6 Subcellular localization of MdHSP90-1、MdHSP90-3、MdHSP90-5 and MdHSP90-11proteins in tobacco
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