TY - JOUR
T1 - Selection and validation of reference genes for normalization of qRT-PCR data to study the cannabinoid pathway genes in industrial hemp
AU - Deguchi, Michihito
AU - Potlakayala, Shobha
AU - Spuhler, Zachary
AU - George, Hannah
AU - Sheri, Vijay
AU - Agili, Ruba
AU - Patel, Aayushi
AU - Rudrabhatla, Sairam
N1 - Publisher Copyright:
© 2021 Deguchi et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
PY - 2021/12
Y1 - 2021/12
N2 - There has been significant interest in researching the pharmaceutical applications of Industrial hemp since its legalization three years ago. The crop is mostly dioecious and known for its production of phytocannabinoids, flavonoids, and terpenes. Although many scientific reports have showed gene expression analysis of hemp through OMICs approaches, unreliable reference genes for normalization of qRT-PCR data make it difficult to validate the OMICs data. Four software packages: GeNorm, NormFinder, BestKeeper, and RefFinder were used to evaluate the differential gene expression patterns of 13 candidate reference genes under osmotic, heavy metal, hormonal, and UV stresses. EF-1α ranked as the most stable reference gene across all stresses, TUB was the most stable under osmotic stress, and TATA was the most stable under both heavy metal stress and hormonal stimuli. The expression patterns of two cannabinoid pathway genes, AAE1 and CBDAS, were used to validate the reliability of the selected reference genes. This work provides useful information for gene expression characterization in hemp and future research in the synthesis, transport, and accumulation of secondary metabolites.
AB - There has been significant interest in researching the pharmaceutical applications of Industrial hemp since its legalization three years ago. The crop is mostly dioecious and known for its production of phytocannabinoids, flavonoids, and terpenes. Although many scientific reports have showed gene expression analysis of hemp through OMICs approaches, unreliable reference genes for normalization of qRT-PCR data make it difficult to validate the OMICs data. Four software packages: GeNorm, NormFinder, BestKeeper, and RefFinder were used to evaluate the differential gene expression patterns of 13 candidate reference genes under osmotic, heavy metal, hormonal, and UV stresses. EF-1α ranked as the most stable reference gene across all stresses, TUB was the most stable under osmotic stress, and TATA was the most stable under both heavy metal stress and hormonal stimuli. The expression patterns of two cannabinoid pathway genes, AAE1 and CBDAS, were used to validate the reliability of the selected reference genes. This work provides useful information for gene expression characterization in hemp and future research in the synthesis, transport, and accumulation of secondary metabolites.
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U2 - 10.1371/journal.pone.0260660
DO - 10.1371/journal.pone.0260660
M3 - Article
C2 - 34928958
AN - SCOPUS:85122012726
SN - 1932-6203
VL - 16
JO - PloS one
JF - PloS one
IS - 12 December
M1 - e0260660
ER -