Cloning and Characterization of a Flavonol Synthase Gene From Litchi chinensis and Its Variation Among Litchi Cultivars With Different Fruit Maturation Periods.

Wei Liu,Zhidan Xiao,Xiangchun Meng,Nonghui Jiang,Chao Fan,Xu Xiang

doi:10.3389/fpls.2018.00567

Abstract

Litchi (Litchi chinensis) is an important subtropical fruit tree with high commercial value. However, the short and centralized fruit maturation period of litchi cultivars represents a bottleneck for litchi production. Therefore, the development of novel cultivars with extremely early fruit maturation period is critical. Previously, we showed that the genotypes of extremely early-maturing (EEM), early-maturing (EM), and middle-to-late-maturing (MLM) cultivars at a specific locus SNP51 (substitution type C/T) were consistent with their respective genetic background at the whole-genome level; a homozygous C/C genotype at SNP51 systematically differentiated EEM cultivars from others. The litchi gene on which SNP51 was located was annotated as flavonol synthase (FLS), which catalyzes the formation of flavonols. Here, we further elucidate the variation of the FLS gene from L. chinensis (LcFLS) among EEM, EM, and MLM cultivars. EEM cultivars with a homozygous C/C genotype at SNP51 all contained the same 2,199-bp sequence of the LcFLS gene. For MLM cultivars with a homozygous T/T genotype at SNP51, the sequence lengths of the LcFLS gene were 2,202–2,222 bp. EM cultivars with heterozygous C/T genotypes at SNP51 contained two different alleles of the LcFLS gene: a 2,199-bp sequence identical to that in EEM cultivars and a 2,205-bp sequence identical to that in MLM cultivar ‘Heiye.’ Moreover, the coding regions of LcFLS genes of other MLM cultivars were almost identical to that of ‘Heiye.’ Therefore, the LcFLS gene coding region may be used as a source of diagnostic SNP markers to discriminate or identify genotypes with the EEM trait. The expression pattern of the LcFLS gene and accumulation pattern of flavonol from EEM, EM, and MLM cultivars were analyzed and compared using quantitative real-time PCR (qRT-PCR) and high-performance liquid chromatography (HPLC) for mature leaves, flower buds, and fruits, 15, 30, 45, and 60 days after anthesis. Flavonol content and LcFLS gene expression levels were positively correlated in all three cultivars: both decreased from the EEM to MLM cultivars, with moderate levels in the EM cultivars. LcFLS gene function could be further analyzed to elucidate its correlation with phenotype variation among litchi cultivars with different fruit maturation periods.

Highlights

Litchi (Litchi chinensis Sonn.), a member of the family Sapindaceae, is an important subtropical fruit tree with high commercial value because of the excellent taste and rich nutritional value of its fruits (Li et al, 2013)
The full-length cDNA sequence of the LcFLS gene amplified from the representative extremely early-maturing (EEM) cultivar ‘Sanyuehong’ was 1,079 bp long and contained a 1,008-bp open reading frame (ORF)
We found a perfect correlation between litchi phylogenetic relationships and the fruit maturation period, based on 90 single nucleotide polymorphisms (SNPs) evenly spaced across the litchi genome (Liu et al, 2015)

Summary

Introduction

Litchi (Litchi chinensis Sonn.), a member of the family Sapindaceae, is an important subtropical fruit tree with high commercial value because of the excellent taste and rich nutritional value of its fruits (Li et al, 2013). Most litchi cultivars ripen between mid-June and mid-July. This results in a short and centralized fruit maturation period, which represents a bottleneck for litchi production in China (Li, 2008). The fruit maturation period of litchi is determined jointly by the flowering time and the fruit development process. Zhao et al (2011) detected five quantitative trait loci for the fruit maturation period using an F1 hybrid population from a cross of litchi cultivars: ‘Maguili’ × ‘Jiaohesanyuehong.’ Ding et al (2015) showed that LcFT1 played an essential part in litchi floral induction, and that sequence differences in the LcFT1 promoter may be one of the causes of the natural variation in the flowering times of different litchi cultivars The molecular mechanisms underlying the fruit maturation period of litchi are not yet fully understood; to our knowledge, there have been only two studies addressing this subject (Zhao et al, 2011; Ding et al, 2015). Zhao et al (2011) detected five quantitative trait loci for the fruit maturation period using an F1 hybrid population from a cross of litchi cultivars: ‘Maguili’ × ‘Jiaohesanyuehong.’ Ding et al (2015) showed that LcFT1 played an essential part in litchi floral induction, and that sequence differences in the LcFT1 promoter may be one of the causes of the natural variation in the flowering times of different litchi cultivars

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