激素处理下马尾松茎干组织qPCR内参基因的筛选

doi:10.3969/j.issn.1674-7968.2022.02.017

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摘要马尾松(Pinus massoniana)的利用价值极高,施加外源激素可促进其茎干的次生生长和木材形成,快速提高生长速度。筛选激素处理下马尾松茎干qPCR实验的最佳内参基因,可为马尾松次生生长的基因表达研究提供技术支持。本研究从马尾松茎干转录组数据库选取14个候选内参基因,利用qPCR技术进行基因表达分析,采用GeNorm、NormFinder和BestKeeper三款软件分析候选内参基因在不同激素—生长素(auxin, IAA)、赤霉素(gibberellin, GA₃)和油菜素内酯(brassinolide, BR)处理条件下马尾松茎干中的表达稳定性,并借助目的基因—纤维素合成酶2 (cellulose synthase 2, CESA2)对筛选出的内参基因进行验证。结果显示,在不同激素处理下,表达稳定性最佳的内参基因为磷酸胆碱胞二苷酰基转移酶1基因(phosphoate cytidine transferase 1, PCYT1)、泛素结合酶E2D基因(ubiquitin-conjugating enzyme E2D, UBE2D)和磷酸葡萄糖变位酶基因(phosphoglucomutase, PGM);目的基因CESA2在激素处理下的表达模式证实上述内参基因组合具有良好的可靠性。本研究筛选的内参基因有助于提高马尾松qPCR实验的准确性和可靠性,为在激素处理下的马尾松基因表达分析提供技术支持。

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	覃慧娟
	范付华
	周紫晶

关键词 ：马尾松, 内参基因, 激素处理, qPCR

Abstract：The utilization value of Pinus massoniana is extremely high, exogenous hormones can promote the secondary growth of its stem and wood formation, and rapidly increase its growth speed. Screening the best internal reference genes for qPCR experiment of P. massoniana stem under hormone treatment can provide technical support for gene expression research of secondary growth of P. massoniana. In this study, 14 candidate internal reference genes were selected from the stem transcriptome database of P. massoniana and gene expression was analyzed by qPCR. The expression stability under different hormone treatment - auxin (IAA), gibberellin (GA₃), and brassinolide (BR) was analyzed by 3 softwares: GeNorm, NormFinder and BestKeeper. The selected internal reference genes were verified by the target gene cellulose synthase 2 (CESA2). The results showed that, under different hormone treatments, the internal reference genes with the best expression stability were phosphoate cytidine transferase 1 (PCYT1), ubiquitin-conjugating enzyme E2D(UBE2D), and phosphoglucomutase (PGM). The expression pattern of target gene CESA2 under hormone treatment confirmed the reliability of the above internal reference gene combination. The internal reference genes screened in this study would be helpful to improve the accuracy and reliability of qPCR experiment in P. massoniana, which provides a solid foundation for the analysis of differential gene expression of P. massoniana under hormone treatment.

Key words： Pinus massoniana Internal reference gene Hormone treatment qPCR

收稿日期: 2021-08-13

ZTFLH:

S718.43

基金资助:贵州省科技计划项目(黔科合平台人才[2018]5261; 黔科合平台人才[2017]5788)

通讯作者: *fhfan1@gzu.edu.cn

引用本文:

覃慧娟, 范付华, 周紫晶. 激素处理下马尾松茎干组织qPCR内参基因的筛选[J]. 农业生物技术学报, 2022, 30(2): 393-401.
QIN Hui-Juan, FAN Fu-Hua, ZHOU Zi-Jing. Screening of qPCR Internal Reference Genes in Stem Tissue of Pinus massoniana Under Hormone Treatment. 农业生物技术学报, 2022, 30(2): 393-401.

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http://journal05.magtech.org.cn/Jwk_ny/CN/10.3969/j.issn.1674-7968.2022.02.017 或 http://journal05.magtech.org.cn/Jwk_ny/CN/Y2022/V30/I2/393

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