Reference gene selection and myosin heavy chain (MyHC) isoform expression in muscle tissues of domestic yak (Bos grunniens)

Wu, Xiaoyun; Zhou, Xiaofang; Ding, Xuezhi; Chen, Min; Liang, Chunnian; Pei, Jie; Xiong, Lin; Bao, Pengjia; Guo, Xian; Yan, Ping

doi:10.1371/journal.pone.0228493

Cited by 14 publications

(3 citation statements)

References 72 publications

(35 reference statements)

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“…In previous studies, YWHAZ has been shown to be a stable transcript during heat stress in the whole blood of sheep (Peletto et al, 2011) and in neutrophils both from healthy sheep and from sheep with foot diseases (Vorachek, Bobe, & Hall, 2013a). Wu et al (2020) have reported that the stability of YWHAZ transcript in different muscles of yaks (Bos grunniens) was higher than that of ACTB, SDHA, B2M, and GAPDH.…”

Section: Resultsmentioning

confidence: 97%

Dietary levels of methionine plus cystine and chelated copper on the chemical composition of eggs and yolk cholesterol of 49-week-old brown laying hens

Sandoval-Lozano

Rondón‐Barragán

2020

Semina: Ciênc. Agrár.

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The Colombian Creole sheep breed has a high economic and social importance for Colombia. Both males and females of this breed are multipurpose animals, and evaluating the production and meat quality of both sexes is important for small farmers in Colombia. This requires the use of tools that help to evaluate critical production points, such as Real-time quantitative polymerase chain reaction (RT-qPCR), which is a widely used molecular tool for the relative quantification of candidate genes in various tissues. For its correct use, the use of housekeeping genes with stable expression, so-called “reference genes”, is required. However, recent studies have shown that the expression of these reference genes can vary among tissues and can be modulated by breed, sex, or external stimuli. Likewise, there is little information regarding the expression of these genes in the Longissimus thoracis et lumborum muscle of male and female Colombian Creole sheep. In this study, the stability in the expression of seven reference genes (ACTB, YWHAZ, SDHA, GAPDH, TUBB2A, B2M, and PGK1) in the Longissimus thoracis et lumborum muscle of male and female Colombian Creole sheep was compared since they are used in RT-qPCR studies to determine the most stable ones for this breed. Twelve animals, six males and six females, with a body weight of 26 ± 4 kg and 12 ± 3 months of age, were used under grazing conditions. Biopsies of the Longissimus thoracis et lumborum muscle were taken, from which RNA was extracted and cDNA was synthesized. Expression was determined using RT-qPCR, and its stability was analyzed by computational algorithms using the geNorm, Normfinder, and BestKeeper software packages, which were integrated using the RefFinder software package. The results indicate that GAPDH, ACTB, and SDHA have the highest stability, whereas the most variable expression was found for B2M. These data provide the basis for more precise results in RT-qPCR studies of gene expression in the muscle of Colombian Creole sheep.

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Section: Resultsmentioning

confidence: 97%

Dietary levels of methionine plus cystine and chelated copper on the chemical composition of eggs and yolk cholesterol of 49-week-old brown laying hens

Sandoval-Lozano

Rondón‐Barragán

2020

Semina: Ciênc. Agrár.

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“…A large proportion of RGs were selected according to previous literature, of which many were housekeeping genes (HKGs) [ 7 , 36 , 37 ]. Moreover, it is widely accepted that the expression levels of HKGs are not stable in some species or tissues, and cannot meet the criteria of RG normalization [ 38 , 39 , 40 ].…”

Section: Discussionmentioning

confidence: 99%

Genome-Wide Identification of Reference Genes for Reverse-Transcription Quantitative PCR in Goat Rumen

Zhao¹,

Cheng²,

Zhang³

et al. 2021

Animals

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As the largest chamber of the ruminant stomach, the rumen not only serves as the principal absorptive surface and nutrient transport pathway from the lumen into the animal, but also plays an important short-chain fatty acid (SCFA) metabolic role in addition to protective functions. Accurate characterization of the gene expression profiles of genes of interest is essential to the exploration of the intrinsic regulatory mechanisms of rumen development in goats. Thus, the selection of suitable reference genes (RGs) is an important prerequisite for real-time quantitative PCR (RT-qPCR). In the present study, 16 candidate RGs were identified from our previous transcriptome sequencing of caprine rumen tissues. The quantitative expressions of the candidate RGs were measured using the RT-qPCR method, and the expression stability of the RGs was assessed using the geNorm, NormFinder, and BestKeeper programs. GeNorm analysis showed that the M values were less than 0.5 for all the RGs except GAPT4, indicating that they were stably expressed in the rumen tissues throughout development. RPS4X and RPS6 were the two most stable RGs. Furthermore, the expressions of two randomly selected target genes (IGF1 and TOP2A), normalized by the selected most stable RGs (RPS4X and RPS6), were consistent with the results of RNA sequencing, while the use of GAPDH and ACTB as RGs resulted in altered profiles. Overall, RPS4X and RPS6 showed the highest expression stability and the lowest coefficients of variation, and could be used as the optimal reference combination for quantifying gene expression in rumen tissues via RT-qPCR analysis.

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“…Specific primer and TaqMan probes (Supplementary Table S1) were designed using species specific gene sequences (Ensemble gene) as described elsewhere [49] and custom-made (LGC Biosearch Technologies, Risskov, Denmark). The NormFinder algorithm-based software [50] was used to determine the best suitable reference gene amongst three previously validated candidates; RPLP0 [51][52][53], TBP [51,54,55] and UTX [52,56,57]. The comparative 2 −(∆∆Ct) method was utilized for graphical representation with all numbers relative to the pre-differentiated state of the non-stored control samples.…”

Section: Quantitative Rt-pcrmentioning

confidence: 99%

Bovine Satellite Cells Isolated after 2 and 5 Days of Tissue Storage Maintain the Proliferative and Myogenic Capacity Needed for Cultured Meat Production

Skrivergaard

Rasmussen

Therkildsen

et al. 2021

IJMS

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Cultured meat is an emerging alternative food technology which aims to deliver a more ethical, sustainable, and healthy muscle-tissue-derived food item compared to conventional meat. As start-up companies are rapidly forming and accelerating this technology, many aspects of this multi-faceted science have still not been investigated in academia. In this study, we investigated if bovine satellite cells with the ability to proliferate and undergo myogenic differentiation could be isolated after extended tissue storage, for the purpose of increasing the practicality for cultured meat production. Proliferation of bovine satellite cells isolated on the day of arrival or after 2 and 5 days of tissue storage were analyzed by metabolic and DNA-based assays, while their myogenic characteristics were investigated using RT-qPCR and immunofluorescence. Extended tissue storage up to 5 days did not negatively affect proliferation nor the ability to undergo fusion and create myosin heavy chain-positive myotubes. The expression patterns of myogenic and muscle-specific genes were also not affected after tissue storage. In fact, the data indicated a positive trend in terms of myogenic potential after tissue storage, although it was non-significant. These results suggest that the timeframe of which viable myogenic satellite cells can be isolated and used for cultured meat production can be greatly extended by proper tissue storage.

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Reference gene selection and myosin heavy chain (MyHC) isoform expression in muscle tissues of domestic yak (Bos grunniens)

Cited by 14 publications

References 72 publications

Dietary levels of methionine plus cystine and chelated copper on the chemical composition of eggs and yolk cholesterol of 49-week-old brown laying hens

Dietary levels of methionine plus cystine and chelated copper on the chemical composition of eggs and yolk cholesterol of 49-week-old brown laying hens

Genome-Wide Identification of Reference Genes for Reverse-Transcription Quantitative PCR in Goat Rumen

Bovine Satellite Cells Isolated after 2 and 5 Days of Tissue Storage Maintain the Proliferative and Myogenic Capacity Needed for Cultured Meat Production

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