Cloning of alfalfa MsMYB86 and analysis of its transcriptional response to abiotic stress

doi:10.11686/cyxb2024416

Abstract

Abstract:

Alfalfa （Medicago sativa） occupies a prominent position in forage production as a high-quality forage with excellent nutritional value and wide adaptability. The MYB family is one of the largest families of transcription factors in plants， and its members play crucial roles in plant growth and development， secondary metabolism， and responses to biotic and abiotic stresses. In this study， MsMYB86 was cloned from alfalfa and its putative encoded protein was analyzed using online websites and software such as ExPASy， Prabi， and SMART. These analyses predicted the relative molecular mass， protein secondary structure， and protein-binding domains of the putative MYB86 protein. Tissue-specific transcript profiles of MsMYB86 and its transcriptional response to different abiotic stresses were analyzed by real-time quantitative polymerase chain reaction （RT-qPCR）. The results showed that the full-length MsMYB86 coding sequence was 1104 bp long， encoding a polypeptide of 367 amino acids. The protein was predicted to have a relative molecular mass of 41.27 kDa， an isoelectric point of 7.10， and a high lipid index of 65.61， indicating that it is a hydrophilic protein. The MsMYB86 protein was predicted to contain two highly conserved SANT-MYB structural domains and to localize in the nucleus. We detected tissue-specific transcript profiles of MsMYB86， and its transcript levels were significantly higher in mature stems than in other tissues. Transcription of the MsMYB86 gene was responsive to drought， salt stress， and abscisic acid treatment， suggesting that it plays a role in the alfalfa response to abiotic stresses. The results of this study provide a theoretical basis for further studies on the role of MsMYB86 in regulating the abiotic stress response of alfalfa.

Key words: alfalfa, MsMYB86, abiotic stress, expression pattern

Ran XIAN, Yu DENG, Qiu-yue FU, Jing-xia JIANG, Jia-li TAO, Tao XU, Hui-sen ZHU, Hui-fang CEN. Cloning of alfalfa MsMYB86 and analysis of its transcriptional response to abiotic stress[J]. Acta Prataculturae Sinica, 2025, 34(9): 162-172.

Figures/Tables 9

Table 1 Primers list

名称Name	上游引物序列Primer-forward sequence （5′-3′）	下游引物序列Primer-reverse sequence （5′-3′）
MsMYB86	ATGGGAAGACATTCTTGCTGCT	CTACATAGTTTGTCCAAAAGCCAC
q-MsMYB86	GCAGCACAATTACCAGGAAGAACAG	CAGAGAGTGGTTTGTGAGTGTTTGG
Actin	CAAAAGATGGCAGATGCTGAGGAT	CATGACACCAGTATGACGAGGTCG

Fig.1 Electrophoresis results of MsMYB86 gene amplification

Fig.2 Alfalfa MsMYB86 CDS full length and coding amino acid sequence

Fig.3 Prediction of structural domain positions of alfalfa MsMYB86 protein （A）， MsMYB86 protein phosphorylation site （B）， MsMYB86 protein hydrophilicity/hydrophobicity （C）， MsMYB86 protein glycosylation site （D）

Fig.4 Subcellular localisation prediction （A） and codon preference analysis （B）

Fig.5 Homologous sequence comparison of MsMYB86 protein in different species

Fig.6 Phylogenetic tree of MYB86 amino acid sequences of different species

Fig.7 Prediction of amino acid secondary structure of alfalfa MsMYB86 （A）， protein interaction network analysis （B）， tertiary structure of alfalfa MsMYB86 protein （C） and tertiary structure of soybean MYB/HD-like transcription factor protein（D）

Fig.8 Plant organs specificity and expression pattern of MsMYB86 gene under abiotic stresses

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