Optimization of reference genes for qRT-PCR analysis of microRNA expression under abiotic stress conditions in sweetpotato

Liu, Xiayu; Liu, Shifang; Zhang, Jie; Wu, Yuhao; Wu, Wan-Yi; Zhang, Yi; Liu, Baoling; Tang, Ruimin; He, Liheng; Li, Runzhi; Jia, Xiaoyun

doi:10.1016/j.plaphy.2020.06.016

Cited by 20 publications

(10 citation statements)

References 37 publications

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“…Additionally, the average Ct values of the candidate RGs ranged from 14.716 (ppt-miR894) to 25.437 (pab-miR395b) ( Figure 2 ). This is similar to the results for sweet potato [ 31 ] and wheat [ 24 ], in which these candidate RGs showed different expression levels with various test materials, with average Ct values from 6.7 to 33.5. These results indicate that in a given sample, no RG has constant expression levels across different conditions; accordingly, it is very important to select suitable RGs for miRNA standardisation under specific experimental conditions.…”

Section: Discussionsupporting

confidence: 87%

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Selection and Optimization of Reference Genes for MicroRNA Expression Normalization by qRT-PCR in Chinese Cedar (Cryptomeria fortunei) under Multiple Stresses

Zhang

Xue

Zhu

et al. 2021

IJMS

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MicroRNA (miRNA) expression analysis is very important for investigating its functions. To date, no research on reference genes (RGs) for miRNAs in gymnosperms, including Cryptomeria fortunei, has been reported. Here, ten miRNAs (i.e., pab-miR159a, cln-miR162, cas-miR166d, pab-miR395b, ppt-miR894, cln-miR6725, novel1, novel6, novel14 and novel16) and three common RGs (U6, 5S and 18S) were selected as candidate RGs. qRT-PCR was used to analyse their expressions in C. fortunei under various experimental conditions, including multiple stresses (cold, heat, drought, salt, abscisic acid and gibberellin) and in various tissues (roots, stems, tender needles, cones and seeds). Four algorithms (delta Ct, geNorm, NormFinder and BestKeeper) were employed to assess the stability of candidate RG expression; the geometric mean and RefFinder program were used to comprehensively evaluate RG stability. According to the results, novel16, cln-miR6725, novel1 and U6 were the most stable RGs for studying C. fortunei miRNA expression. In addition, the expression of three target miRNAs (aly-miR164c-5p, aly-miR168a-5p and smo-miR396) was examined to verify that the selected RGs are suitable for miRNA expression normalisation. This study may aid further investigations of miRNA expression/function in the response of C. fortunei to abiotic stress and provides an important basis for the standardisation of miRNA expression in other gymnosperm species.

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Section: Discussionsupporting

confidence: 87%

“…The most commonly used RGs in plant miRNA qRT-PCR are still U6 and 5S [ 20 , 21 ]. Previous studies have shown that certain miRNAs have higher stability than mRNAs, as in rice ( Oryza sativa ) [ 30 ], sweet potato ( Ipomoea batatas ) [ 31 ] and peach [ 26 ]. Unfortunately, there are no universal RGs [ 19 ].…”

Section: Discussionmentioning

confidence: 99%

Selection and Optimization of Reference Genes for MicroRNA Expression Normalization by qRT-PCR in Chinese Cedar (Cryptomeria fortunei) under Multiple Stresses

Zhang

Xue

Zhu

et al. 2021

IJMS

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“…Hou et al (2021) used the TRIzol method to isolate RNA and analyze genes from the Bgalactosidase family, which are involved in the response of biotic and abiotic stresses. In addition, the TRIzol method has also been used on the transcriptome sequencing of sweet potato (Tao et al, 2012), and on the characterization of ERD15, a gene that seems to be involved in the defense response to water stress in sweet potato (Shao et al, 2014), to identify water deficit responsive genes in RNAseq and RT-qPCR studies (Arisha et al, 2020), in the validation of reference genes for the expression normalization of miRNAs (Liu et al, 2020), and for the evaluation of genes associated to starch and sucrose metabolism (Ma et al, 2020;Qin et al, 2021).…”

Section: Rna Extraction Methodsmentioning

confidence: 99%

Research Article Evaluation of extraction methods for obtaining high-quality RNA from sweet potato

Gonçalves¹,

Daúde²,

Souza³

et al. 2021

Genet. Mol. Res.

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“…β-tubulin gene was used as housekeeping gene (internal control) to normalize the relative expression of genes. RT-qPCR was performed in CFX96 Touch Deep Well Real Time PCR System (Bio-Rad, USA) with the program in previous study (Liu et al, 2020). Each gene for qPCR was performed by a biological/experimental triplicate.…”

Section: Validation Of Rna-seq Data Using Qpcrmentioning

confidence: 99%

The SPEECHLESS-induced stomatal increase is required for the salt tolerance of oil palm

Song

Wang

Lai

et al. 2021

Preprint

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Oil palm is the most productive oil producing plant. Salt stress leads to growth damage and decrease in yield of oil palm. However, the physiological responses of oil palm to salt stress and their underlying mechanisms are not clear. RNA-Seq for leaf samples from young palms challenged under three levels of salts (100, 250 and 500 mM NaCl) and control for 14 days was conducted. Diverse signalling pathways were involved in responses to different levels of salt stress. All the three levels of salt stress activated EgSPCH expression and induced stomatal density of oil palm, which was contrasting to that in Arabidopsis. Under strong salt stress group, oil palm removed excessive salt via stomata. Overexpression of EgSPCH in Arabidopsis increased the stomatal production but lowered the salt tolerance. These data suggest that in oil palm, salt activates EgSPCH to generate more stomata in response to salt stress. Our results shed a light on the cellular response to salt stress of oil palm and provide new insights into the mechanisms of different salt-induced stomatal development between halophytes and glycophytes.

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Optimization of reference genes for qRT-PCR analysis of microRNA expression under abiotic stress conditions in sweetpotato

Cited by 20 publications

References 37 publications

Selection and Optimization of Reference Genes for MicroRNA Expression Normalization by qRT-PCR in Chinese Cedar (Cryptomeria fortunei) under Multiple Stresses

Selection and Optimization of Reference Genes for MicroRNA Expression Normalization by qRT-PCR in Chinese Cedar (Cryptomeria fortunei) under Multiple Stresses

Research Article Evaluation of extraction methods for obtaining high-quality RNA from sweet potato

The SPEECHLESS-induced stomatal increase is required for the salt tolerance of oil palm

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