Cloning and expression analysis of CiMYB46 in the graft healing process of Carya illinoinensis

MO Zhenghai, LI Fengda, SU Wenchuan, CAO Fan, PENG Fangren, LI Yongrong

JOURNAL OF NANJING FORESTRY UNIVERSITY ›› 2019, Vol. 43 ›› Issue (5) : 156-162.

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JOURNAL OF NANJING FORESTRY UNIVERSITY ›› 2019, Vol. 43 ›› Issue (5) : 156-162. DOI: 10.3969/j.issn.1000-2006.201811041

Cloning and expression analysis of CiMYB46 in the graft healing process of Carya illinoinensis

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Abstract

【Objective】 To explore the molecular mechanism of graft healing in pecan, we cloned the CiMYB46 gene and analyzed its expression pattern and cis-elements. 【Method】 We extracted RNA from graft unions of pecan plants at different developmental stages and designed primers, based on our previous transcriptome results, to clone the desired gene by RT-PCR. In addition, genomic DNA was used as a template for PCR amplification and cloning of the promoter area of the desired gene.【Result】 The amplified fragment obtained by RT-PCR was cloned and sequenced. It contained an open reading frame of 969 bp that started with the initiation codon, ATG and ended with the termination codon, TAA, encoding a polypeptide of 322 amino acids. The deduced amino acid sequence of the cloned gene contained a MYB domain and was closely clustered withArabidopsis AtMYB46. Thus, we designated our sequence as CiMYB46. Real-time PCR analyses demonstrated that CiMYB46 was highly expressed during the vascular formation stage of graft union development in pecan and had a similar expression profile to that of some functional genes involved in secondary cell wall synthesis. A 1 070 bp promoter region upstream of the initiation codon of CiMYB46 was obtained through PCR and cloning, using genomic DNA as a template. PlantCARE analysis revealed that the promoter region of CiMYB46 contained two basic cis-acting elements, a CAAT box and a TATA-box, drought responsive elements, and hormone responsive elements, including ABA, Me-JA, GA, and SA. 【Conclusion】 CiMYB46 might be associated with vascular bundle formation during the graft healing process of pecan. It may be induced by GA.

Key words

Carya illinoinensis / grafting / gene cloning / CiMYB46 / MYB transcription factor

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MO Zhenghai , LI Fengda , SU Wenchuan , et al . Cloning and expression analysis of CiMYB46 in the graft healing process of Carya illinoinensis[J]. JOURNAL OF NANJING FORESTRY UNIVERSITY. 2019, 43(5): 156-162 https://doi.org/10.3969/j.issn.1000-2006.201811041

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