Sequence and Tissue Expression Analysis of Monosaccharide Transporter Gene ShSTP7 in Sugarcane

doi:10.13560/j.cnki.biotech.bull.1985.2021-1537

Abstract

Abstract:

Sugar transporter protein（ST）genes play important roles on yield formation and sugar accumulation in sugarcane（Saccharum spp. hybrids）. The function analysis of ST genes facilitates to clarify the sugar accumulation mechanism and sugarcane genetic improvement. The ST gene ShSTP7 was isolated from sugarcane,and its biological characteristics,subcellular location and expression pattern of ShSTP7 were analyzed by bioinformatics software and techniques including RT-PCR,real-time PCR and protoplast subcellular location. The results showed that a sugar transporter gene ShSTP7 cDNA was isolated from sugarcane leaves. This gene was 1 619 bp length containing 1 557 bp open reading frame and encoded 518 amino acids. ShSTP7-encoded protein ShSTP7 belonged to facilitated diffusion superfamily and sugar transporter family. It had 12 transmembrane structures and sugar transport conserve domain. ShSTP7 was highly homologous with STP7 from S. spontaneum,Zay mays and Sorghum bicolor. The subcellular location analysis indicated that the green fluorescence signal of ShSTP7-GFP fusion protein appeared in the region of plasma membrane. The expression analysis of ShSTP7 demonstrated that it mainly expressed in sugarcane leaves and less expressed in sugarcane stalks. The expression of ShSTP7 in immature internodes 1-3 of high-sugar-content sugarcane cultivar was higher than that of low-sugar-content sugarcane cultivar. However,the expression of ShSTP7 in the leaves and internodes 4-24 of high-sugar-content sugarcane cultivar was lower than that of low-sugar-content sugarcane cultivar. These results indicate that ShSTP7 is a sugar transporter on plasma membrane uptaking sugars from apoplasmic space and plays an important role on sugar transport in sugarcane leaves,which lays a foundation in clarifying the gene function of monosaccharide transporters and its application.

Key words: monosaccharide transporter protein, ShSTP7, sugar uptake, expression analysis, subcellular location, Saccharum spp. hybrids

ZHAO Ting-ting, WANG Jun-gang, WANG Wen-zhi, FENG Cui-lian, FENG Xiao-yan, ZHANG Shu-zhen. Sequence and Tissue Expression Analysis of Monosaccharide Transporter Gene ShSTP7 in Sugarcane[J]. Biotechnology Bulletin, 2022, 38(4): 72-78.

Figures/Tables 5

Table 1 Design and application of primers

引物名称 Primer name	引物序列 Primer sequence（5'-3'）	引物用途 Application of primers
ShSTP7-F	TGGTAGTGGCTCGTCTTGGAG	克隆引物
ShSTP7-R	TCTTCACTTGTGATTGTTGCTGG	克隆引物
ShSTP7Q-F	GGACCCATTTCTGGAGAA	荧光定量引物
ShSTP7Q-R	TTGCAGTAGTTGTTCTTCC	荧光定量引物
GADPH-F	CACGGCCACTGGAAGCA	荧光定量引物
GADPH-R	TCCTCAGGGTTCCTGATGCC	荧光定量引物
ShSTP7G-F	ACTAGTATGGCGGGCGGCGG- GATG	载体构建引物
ShSTP7G-R	ACTAGTCTTGTGATTGTTGC- TGGCAGGT	载体构建引物

Fig.1 PCR amplification electrophoresis of sugarcane ShSTP7 cDNA

Fig.2 Amino acid sequence homology（A）,transmembrane structure（B）and phylogenetic tree analysis（C）of sugarcane ShSTP7 A：Amino acid sequence homology of ShSTP7. B：Transmembrane structure analysis of ShSTP7. C：Phylogenetic tree analysis of ShSTP7

Fig.3 Expression pattern analysis of ShSTP7 in different sugarcane cultivars A：Expression pattern of ShSTP7 in ROC22. B：Expression pattern of ShSTP7 in 28NG251（HS）and MUCK CHE（LS）. YL：Immature leave. ML：Mature leave. J1：Internode 1-2. J2：Internode 4-5. J3：Internode 7-8. J4：Internode 10-11. J5：Internode 12-13. J6：Internode 15-16. J7：Internode 18-19. J8：Internode 23-24

Fig.4 Subcellular location result of ShSTP7 -GFP fusion protein in protoplast of rice mesophyll cells A：Enzyme digestion by Spe I of ShSTP7-GFP fusion expression construct. B：Subcellular locations of ShSTP7-GFP and pCAMBIA1302-GFP in protoplast of rice mesophyll cells. From left to right, images are fluorescence and bright merged field, fluorescence field and bright field, respectively

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