Análise e validação de polimorfismo de nucleotídeo único (SNP) na região codificadora de duas cultivares contrastantes da cana-de-açúcar submetidas à prolongada limitação hídrica
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Data
2022-11-25
Autores
Carvalho, Bárbara Balisa
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Universidade Estadual Paulista (Unesp)
Resumo
A cana-de-açúcar é uma cultura de grande importância econômica. O estresse hidrico é um dos fatores que prejudica a produtividade e tem sido alvo de preocupação por parte dos melhoristas e da comunidade científica frente ao cenário de mudanças climáticas. As ferramentas moleculares são uma forte aliada aos programas de melhoramento no desenvolvimento de cultivares resilientes às mudanças climáticas. O objetivo do estudo foi validar o nível de expressão gênica de três transcritos de cana-de-açúcar e comparar o perfil da expressão destes transcritos com SNPs exclusivos na cultivar tolerante com os da cultivar sensível sem SNPs, através da técnica de qRT-PCR. Foram utilizadas duas cultivares de cana-de-açúcar, SP81-3250, considerada padrão para tolerância, e RB855453, padrão para cultivar sensível, as quais foram submetidas ao estresse hídrico prolongado (90 dias). A coleta de folhas foi realizada aos 30, 60 e 90 dias após o início do déficit hídrico e as amostras de RNA obtidas foram submetidas ao sequenciamento (RNA-seq). A partir do transcriptoma montado das duas cultivares juntas, foram identificados SNPs na região codificadora de genes relacionados à tolerância ao estresse hídrico, exclusivos da cultivar tolerante. Os transcritos escolhidos para a análise foram também diferencialmente expressos pela técnica do RNA-seq: Serina/Treonina Quinase, um fator de transcrição ABF e um transcrito que codifica uma Proteína Universal do Estresse (USP). Em geral, foi visto uma diferença significativa na expressão dos genes na cultivar tolerante com SNPs em comparação com a sensível sem SNPs. A cultivar tolerante percebe o estresse primeiro e, assim, ativa mais rapidamente a maquinária gênica responsável pelo mecanismo de proteção contra a perda de água, o que afeta também o sistema enzimático antioxidante protetor, diminuindo a perda de água desta cultivar. A presença de SNP na cultivar tolerante pode ter influência na maior indução dos transcritos analisados em comparação com a cultivar sensível, podendo estar ligados à forma em que a cultivar lida com esse estresse, porém é necessário estudos moleculares mais aprofundados sobre o assunto.
- Sugarcane is a crop of great economic importance. Water stress is one of the factors that impair productivity and has been the subject of concern by breeders and the scientific community in the face of climate change. Molecular tools are a strong ally to breeding programs in the development of cultivars that are resilient to climate change. The objective of the study was to validate the gene expression level of 3 sugarcane transcripts and compare the expression profile of these transcripts with exclusive SNPs in the tolerant cultivar with those of the sensitive cultivar without SNPs, using the qRT-PCR technique. Two sugarcane cultivars were used, SP81-3250, considered standard for tolerance and RB855453, standard for sensitive cultivar, which were subjected to prolonged water stress (90 days). Leaf collection was performed at 30, 60 and 90 days after the onset of water deficit and the RNA samples obtained were submitted to sequencing (RNA-seq). From the assembled transcriptome of the two cultivars together, SNPs were identified in the coding region of genes related to tolerance to water stress, exclusive to the tolerant cultivar. The transcripts chosen for analysis were also differentially expressed by the RNA-seq technique: Serine/Threonine Kinase, an ABF transcription factor and a transcript that encodes a Universal Stress Protein (USP). Overall, a significant difference in gene expression was seen in the SNP-tolerant cultivar compared to the non-SNP-sensitive cultivar. The tolerant cultivar perceives stress first and, therefore, more quickly activates the genetic machinery responsible for the protection mechanism against water loss, which also affects the protective antioxidant enzymatic system, reducing the water loss of this cultivar. The presence of SNP in the tolerant cultivar may have an influence on the greater induction of the analyzed transcripts compared to the sensitive cultivar, and may be linked to the way in which the cultivar deals with this stress, but further molecular studies are needed on the subject.
- Sugarcane is a crop of great economic importance. Water stress is one of the factors that impair productivity and has been the subject of concern by breeders and the scientific community in the face of climate change. Molecular tools are a strong ally to breeding programs in the development of cultivars that are resilient to climate change. The objective of the study was to validate the gene expression level of 3 sugarcane transcripts and compare the expression profile of these transcripts with exclusive SNPs in the tolerant cultivar with those of the sensitive cultivar without SNPs, using the qRT-PCR technique. Two sugarcane cultivars were used, SP81-3250, considered standard for tolerance and RB855453, standard for sensitive cultivar, which were subjected to prolonged water stress (90 days). Leaf collection was performed at 30, 60 and 90 days after the onset of water deficit and the RNA samples obtained were submitted to sequencing (RNA-seq). From the assembled transcriptome of the two cultivars together, SNPs were identified in the coding region of genes related to tolerance to water stress, exclusive to the tolerant cultivar. The transcripts chosen for analysis were also differentially expressed by the RNA-seq technique: Serine/Threonine Kinase, an ABF transcription factor and a transcript that encodes a Universal Stress Protein (USP). Overall, a significant difference in gene expression was seen in the SNP-tolerant cultivar compared to the non-SNP-sensitive cultivar. The tolerant cultivar perceives stress first and, therefore, more quickly activates the genetic machinery responsible for the protection mechanism against water loss, which also affects the protective antioxidant enzymatic system, reducing the water loss of this cultivar. The presence of SNP in the tolerant cultivar may have an influence on the greater induction of the analyzed transcripts compared to the sensitive cultivar, and may be linked to the way in which the cultivar deals with this stress, but further molecular studies are needed on the subject.
Descrição
Palavras-chave
Transcriptoma, qRT-PCR, Poliploides, Estresse Hídrico, Saccharum spp., Cana-de-açucar