Scientia Agricultura Sinica ›› 2012, Vol. 45 ›› Issue (23): 4759-4769.doi: 10.3864/j.issn.0578-1752.2012.23.002

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Cloning and Expression Analysis of A Putative B Class MADS-box Gene of AcPI in Onion

 LI  Hong-You, WANG  Chan, LI  Li-Lin, WANG  Yong-Qin, ZHAO  Rui   

  1. 1.College of Horticulture, Shenyang Agricultural University, Shenyang 110161;
    2.Beijing Vegetable Research Center, Beijing Academy of Agriculture and Forestry Sciences/Key Laboratory of Biology and Genetic Improvement of Horticultural Crops (North China), Ministry of Agriculture, Beijing 100097;
    3.School of Life Science, Shanxi University, Taiyuan 030005
  • Received:2012-08-15 Online:2012-12-01 Published:2012-09-14

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Abstract: 【Objective】The aim of this study is to clone the PI/GLO subfamily MADS-box gene of class B of the floral identity in onion, to analyze its sequence characteristics and the temporal and spatial expression pattern. The results will provide a foundation for further dissection of the molecular and genetics mechanisms of floral organ development in onion in detail.【Method】Using the total RNA from the floral buds of onion (Allium cepa) as the template, we design the degenerate primers based on homology cloning strategy, and then the full-length cDNA sequence of AcPI was obtained through a combined reverse transcription-PCR (RT-PCR) and RACE techniques strategy. The bioinformatics methods were used to analyze the sequence characteristics of this gene and the combination of RT-PCR and Real-time PCR methods were used to investigate the temporal and spatial expression pattern of AcPI gene.【Result】The cloned full-length cDNA of AcPI was 931 bp in length, containing a 615 bp open reading frame (ORF) which encodes 205 amino acids. Protein alignment analysis indicated that AcPI protein had typical structure domains of plants MADS-box protein, containing a MADS domain and a K domain. Protein sequence identity search showed that AcPI is similar to NTPI (Narcissus tazetta var. chinensis), HoMADS2 (Hyacinthus orientalis) and GLO (Antirrhinum majus) with similarities of 78%, 75% and 52% respectively. Phylogenetic tree reconstructed clearly showed that AcPI protein was classed into PI subgroup of B class MADS-box protein family. RT-PCR analysis indicated that expression of the AcPI gene was restricted to the flowers as its mRNA was not detected in vegetative tissues such as roots, stems, leaves. Real-time PCR analysis showed that AcPI gene mainly expressed in the first, second and third whorls of flower organ and only weak expression was detected in the fourth whorls of flower organ in all three flower bud developmental stages. Furthermore, the expression level of AcPI was increasing in all flower organs except in the first whorl flower organ is first increases and then decreases in the throughout growing process of flower buds.【Conclusion】The extended expression of AcPI in the first whorl floral organ supported the modified ABC model by van Tunen, but it also expressed in the fourth whorl floral organ, which showed that AcPI gene may play an important role in the development of carpels.

Key words: onion , floral organs , MADS-box gene , gene cloning , expression analysis

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