Molecular cloning and expression of an encoding galactinol synthase gene (AnGolS1) in seedling of Ammopiptanthus nanus.

Yudong Liu,Yi He,Li Zhang,Lijing Chen,Mingfang Qi,Yanye Ruan,Tao Xu,Hui Ma,Chuanqiang Xu

doi:10.1038/srep36113

Abstract

Based on the galactinol synthase (AnGolS1) fragment sequence from a cold-induced Suppression Subtractive Hybridization (SSH) library derived from Ammopiptanthus nanus (A. nanus) seedlings, AnGolS1 mRNA (including the 5′ UTR and 3′ UTR) (GenBank accession number: GU942748) was isolated and characterized by rapid amplification of cDNA ends polymerase chain reaction (RACE–PCR). A substrate reaction test revealed that AnGolS1 possessed galactinol synthase activity in vitro and could potentially be an early-responsive gene. Furthermore, quantitative real-time PCR (qRT-PCR) indicated that AnGolS1 was responded to cold, salts and drought stresses, however, significantly up-regulated in all origans by low temperatures, especially in plant stems. In addition, the hybridization signals in the fascicular cambium were strongest in all cells under low temperature. Thus, we propose that AnGolS1 plays critical roles in A. nanus low-temperature stress resistance and that fascicular cambium cells could be involved in AnGolS1 mRNA transcription, galactinol transportation and coordination under low-temperature stress.

Highlights

Based on the galactinol synthase (AnGolS1) fragment sequence from a cold-induced Suppression Subtractive Hybridization (SSH) library derived from Ammopiptanthus nanus (A. nanus) seedlings, AnGolS1 mRNA (GenBank accession number: GU942748) was isolated and characterized by rapid amplification of cDNA ends polymerase chain reaction (RACE–PCR)
An AnGolS1 fragment was obtained from a cold-induced SSH library derived from A. nanus seedlings, and the 5′-end and 3′-end sequences were amplified by 5′- and 3′-RACE, respectively
AnGolS1 shares a high degree of homology with a variety of GolS proteins from A. mongolicus (AmGolS, 98%), Glycine max (GmGolS1, 85.80%), Medicago truncatula (MtGolS, 82.32%), Nicotiana tabacum (NtGolS2, 76.35%), and A. thaliana (AtGolS1, 73.26%) (Fig. 1)

Summary

Introduction

Based on the galactinol synthase (AnGolS1) fragment sequence from a cold-induced Suppression Subtractive Hybridization (SSH) library derived from Ammopiptanthus nanus (A. nanus) seedlings, AnGolS1 mRNA (including the 5′ UTR and 3′ UTR) (GenBank accession number: GU942748) was isolated and characterized by rapid amplification of cDNA ends polymerase chain reaction (RACE–PCR). Seeds accumulate an abundance of RFOs, and GolS activity gradually increases before stachyose rises, reaching its peak after flowering and decreasing to a plateau in dry seeds[13,23,24,25]. Fleshy fruits such as melons actively synthesize raffinose and stachyose, but the contents of both sugars are very low, thereby suggesting that RFOs are rapidly catabolized[26,27]. The temperature varies dramatically from −30 °C to 47.6 °C; at the same time, the climate is extremely arid because annual precipitation is lower than 20 mm, whereas annual evaporation is higher than 2,000 mm[29,30]

Methods

Results

Conclusion