| 谭健俊,黄李珊,周熠玮,等.姜荷花苞片实时荧光定量PCR内参基因筛选[J].广东农业科学,2024,(12-2501):- |
|
PDF
HTML
姜荷花苞片实时荧光定量PCR内参基因筛选 |
| Screening of Reference Genes for Quantitative Real-time PCR in Curcuma alismatifolia Bracts |
| 投稿时间：2024-09-26 修订日期：2024-10-25 |
| DOI： |
| 中文关键词: 实时荧光定量PCR 姜荷花 内参基因 苞片颜色 MDH；基因表达模式 |
| 英文关键词: qRT-PCR Curcuma alismatifolia reference gene bract color MDH the expression patterns of genes |
| 基金项目:广东省基础与应用基础研究基金(2022A1515110580)；国家自然科学基金（32401661） |
|
| 摘要点击次数: 19 |
| 全文下载次数: 0 |
| 中文摘要: |
| 【目的】实时荧光定量PCR（Quantitative Real-time PCR，qRT-PCR）是检测特定基因在植物组织内表达水平高低最简便高效方法，筛选稳定的内参基因可以确保其结果可靠准确。迄今为止，尚未有过姜荷花内参基因研究的报道，筛选适用于姜荷花苞片组织qRT-PCR分析的内参基因，可为苞片颜色相关基因功能研究奠定理论基础。【方法】以9个不同颜色的姜荷花品种为试材，根据姜荷花基因组数据选择常用的内参基因，包括甘油醛-3-磷酸脱氢酶（GAPDH）、18S核糖体RNA（18S rRNA）、PP2A（蛋白磷酸酶2A）、泛素蛋白（UBQ）、苹果酸脱氢酶（MDH）、肌动蛋白（Actin）、微管蛋白（TUB）和苯丙氨酸解氨酶（PAL）8个看家基因，使用ΔCt、geNorm、NormFinder和BestKeeper 4种方法分析评估基因表达的稳定性，通过RefFinder程序综合评价筛选出姜荷花苞片组织的最佳内参基因。【结果】分析结果表明，8个内参基因均扩增出单一主峰，引物标准曲线相关系数R2 ≥ 0.98，扩增效率为99.5%~125.0%。利用RefFinde程序综合评价得出候选内参基因稳定性综合排名，高低排序依次为MDH＞Actin＞TUB＞18S rRNA＞PP2A＞GAPDH＞UBQ＞PAL，表明不同品种姜荷花苞片最佳内参基因是MDH，其次为Actin和TUB，PAL稳定性最差，不适合作为姜荷花苞片组织的内参基因。【结论】MDH作为不同品种姜荷花苞片组织的稳定内参基因，为后续深入开展姜荷花苞片颜色合成相关基因的表达模式分析提供参考。 |
| 英文摘要: |
| 【Objective】 The quantitative real-time PCR is the most direct and efficient technique for quantifying gene expression levels in plant tissues, ensuring that the selection of stable reference genes is crucial for obtaining reliable and precise results. To date, the research of reference genes has not been documented in Curcuma alismatifolia. Identification of suitable reference genes for qRT-PCR analysis in C. alismatifolia bract will establishe the theoretical foundation for investigating the functional genes associated with bract color. 【Method】 Nine C. alismatifolia cultivars were employed in this experiment. Based on the genomic information, eight housekeeping genes including glyceraldehyde-3-phosphate dehydrogenase (GAPDH), 18S rRNA, protein phosphatase 2A (PP2A), ubiquitin (UBQ), malic dehydrogenase (MDH), actin 1 (Actin), β-tubulin (TUB) and phenylalanine ammonia lyase (PAL) were selected, and four different calculation programs including ΔCt, geNorm, NormFinder and BestKeeper were used to evaluate the expression stability of 8 genes. The RefFinder program was utilized to comprehensively assess the optimal reference genes for C. alismatifolia bract tissue. 【Result】 Results indicated that all the eight reference genes amplified a single band, and the dissolution curves were all single peak with R2 ≥ 0.98 and amplification efficiency value of 99.5%~125.0%. The comprehensive ranking of candidate reference genes stability was obtained using the RefFinder program, with the order from high to low being MDH＞Actin＞TUB＞18S rRNA＞PP2A＞GAPDH＞UBQ＞PAL. The most suitable reference genes for different cultivars of C. alismatifolia bract were MDH, followed by Actin and TUB. The stability of PAL is the least favorable, rendering it unsuitable as a reference gene for C. alismatifolia bract tissue. 【Conclusion】 The stable expression of MDH in different cultivars of C. alismatifolia bract serves as a reliable reference gene for further analysis of the expression patterns of genes involved in color synthesis in C. alismatifolia. |
|
查看/发表评论 下载PDF阅读器 |
