Validation of reference genes for normalization of gene expression by qRT-PCR in a resveratrol-producing entophytic fungus (Alternaria sp. MG1).
Validation of reference genes for normalization of gene expression by qRT-PCR in a resveratrol-producing entophytic fungus (Alternaria sp. MG1).
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DOI：
10.1186/s13568-016-0283-z
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发表时间：
2016-12
期刊：
影响因子：
3.7
通讯作者：
Liu YL
中科院分区：
文献类型：
--
作者：
Che JX;Shi JL;Lu Y;Liu YL
分离自葡萄属葡萄的内生真菌链格孢菌MG 1能独立产生白藜芦醇，表明该菌株含有白藜芦醇生物合成的关键基因。这些关键基因的鉴定对于了解该菌株中白藜芦醇生物合成途径至关重要，这是目前微生物中未知的。qRT-PCR是一种在基因表达水平上鉴定与未知通路相关的关键基因的有效和广泛使用的方法。该菌株中稳定参考基因的验证对于qRT-PCR数据标准化至关重要，尽管已报告了其他链格孢属菌株的结果。在这项研究中，九个候选参考基因，包括TUBA，EF 1，EF 2，UBC，UFD，RPS 5，RPS 24，ACTB和18 S的表达稳定性进行了评估，在一组不同的六个样品代表不同的生长期。我们比较了细胞培养条件和白藜芦醇生产的优化条件。使用四种统计软件对结果进行比较。TUBA和EF 1的组合被发现适合于在不同发育阶段对链格孢属MG 1进行标准化，并且发现18 S最不稳定。本研究验证的参考基因为进一步研究链格孢菌MG 1的基因表达和分子机制以及提高其次生代谢产物产量奠定了基础。据我们所知，这是第一次验证链格孢属中具有产生白藜芦醇能力的参考基因。此外，这些结果为其他链格孢属物种参考基因的选择提供了有用的指导。本文的在线版本（doi：10.1186/s13568-016-0283-z）包含补充材料，可供授权用户使用。
Alternaria sp. MG1, an endophytic fungus isolated from Vitis vinifera, can independently produce resveratrol, indicating that this species contains the key genes for resveratrol biosynthesis. Identification of these key genes is essential to understand the resveratrol biosynthesis pathway in this strain, which is currently unknown in microorganisms. qRT-PCR is an efficient and widely used method to identify the key genes related to unknown pathways at the level of gene expression. Verification of stable reference genes in this strain is essential for qRT-PCR data normalization, although results have been reported for other Alternaria sp. strains. In this study, nine candidate reference genes including TUBA, EF1, EF2, UBC, UFD, RPS5, RPS24, ACTB and 18S were evaluated for expression stability in a diverse set of six samples representing different growth periods. We compared cell culture conditions and an optimized condition for resveratrol production. The comparison of the results was performed using four statistical softwares. A combination of TUBA and EF1 was found to be suitable for normalization of Alternaria sp. MG1 in different developmental stages, and 18S was found to be the least stable. The reference genes verified in this study will facilitate further research to explore gene expression and molecular mechanisms as well as the improvement of secondary metabolite yields in Alternaria sp. MG1. To our knowledge, this is the first validation of reference genes in Alternaria with the capability to produce resveratrol. Additionally, these results provide useful guidelines for the selection of reference genes in other Alternaria species. The online version of this article (doi:10.1186/s13568-016-0283-z) contains supplementary material, which is available to authorized users.
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影响因子：
9.5
作者：
Marabita F;de Candia P;Torri A;Tegnér J;Abrignani S;Rossi RL
通讯作者：
Rossi RL
DOI：
10.1073/pnas.1011289108
发表时间：
2011-02-08
影响因子：
11.1
作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
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影响因子：
3.3
作者：
Kong, Fanna;Cao, Min;Mao, Yunxiang
通讯作者：
Mao, Yunxiang
影响因子：
5.3
作者：
Exposito-Rodriguez, Marino;Borges, Andres A.;Borges-Perez, Andres;Perez, Jose A.
通讯作者：
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