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[内生真菌对大花红景天红景天苷生物合成途径中关键酶基因表达的影响]

[Effect of endophytic fungus on expression of key enzyme genes in pathway of salidroside biosynthesis in Rhodiola crenulata].

作者信息

Wang Meng-liang, Wang Ya-nan, Cui Jin-long, Wang Jun-hong

出版信息

Yao Xue Xue Bao. 2016 Dec;51(12):1920-5.

Abstract

One strain of endophytic fungus ZPRa-R-1 was obtained for the capacity of promoting production of salidroside in Rhodiola crenulata. To explain the mechanism of salidroside biosynthesis in host plant, eight housekeeping genes were evaluated, and the evaluation method was created for the expression activities of four key enzyme genes PAL (phenylalanine ammonia-lyase), TyDC (tyrosine decarboxylase), TAT (tyrosine transaminase), UDPGT (UDP-glucosyltransferase) referenced double reference genes in biosynthesis pathway of salidroside in R. crenulata. Stabilities of housekeeping genes were confirmed by real-time fluorescent quantitative PCR technology and three softwares including geNorm, NormFinder and BestKeeper, then relative expressions of key enzyme genes were analysized by the 2-ΔΔCt method. The results showed that the most stable gene was GAPDH, followed by PCS, and the most appropriate reference of internal genes were combination with two genes in R. crenulata inoculated with endophytic fungus ZPRa-R-1. Under symbiosis conditions, regularity changes of key enzyme genes affected by endophytic fungus ZPRa-R-1 were as follows: the relative expression activity of PAL attached to peak value, which was 4.9 times as that of control group when inoculated ten days. The relative expression of TyDC reached the maximum value, which was 2.8 times of that control after inoculating 12 days. The relative expression of UDPGT actually reach 17.1 times than that of control after inoculating 8 days. However, the relative expression of TAT was not affected by this fungus. The changes of four key enzyme genes are positively correlated with the changes of salidroside content in R. crenulata.

摘要

从西藏红景天中筛选获得1株具有促进红景天苷合成能力的内生真菌ZPRa-R-1。为阐明红景天苷在宿主植物中的生物合成机制,对8个管家基因进行评估,并建立了以双内参基因法评估西藏红景天红景天苷生物合成途径中4个关键酶基因苯丙氨酸解氨酶(PAL)、酪氨酸脱羧酶(TyDC)、酪氨酸转氨酶(TAT)、尿苷二磷酸葡萄糖基转移酶(UDPGT)表达活性的方法。采用实时荧光定量PCR技术结合geNorm、NormFinder和BestKeeper 3种软件对管家基因的稳定性进行验证,然后用2-ΔΔCt法分析关键酶基因的相对表达量。结果表明,最稳定的基因是GAPDH,其次是PCS,在接种内生真菌ZPRa-R-1的西藏红景天中,最适宜的内参基因组合是2个基因。共生条件下,内生真菌ZPRa-R-1影响关键酶基因的变化规律如下:PAL的相对表达活性在接种第10天达到峰值,是对照组的4.9倍;TyDC的相对表达量在接种第12天达到最大值,是对照组的2.8倍;UDPGT的相对表达量在接种第8天达到最大值,是对照组的17.1倍;而TAT的相对表达量不受该内生真菌的影响。4个关键酶基因的变化与西藏红景天中红景天苷含量的变化呈正相关。

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