Sun Rui, Chang Yuansheng, Yang Fengqiu, Wang Yi, Li Hui, Zhao Yongbo, Chen Dongmei, Wu Ting, Zhang Xinzhong, Han Zhenhai
Institute for Horticultural Plants, College of Agronomy and Biotechnology, China Agricultural University, Beijing, 100193, China.
Changli Institute for Pomology, Hebei Academy of Agricultural and Forestry Science, Changli, Heibei 066600, China.
BMC Genomics. 2015 Oct 5;16:747. doi: 10.1186/s12864-015-1946-x.
Genetic map based quantitative trait locus (QTL) analysis is an important method for studying important horticultural traits in apple. To facilitate molecular breeding studies of fruit quality traits in apple, we aim to construct a high density map which was efficient for QTL mapping and possible to search for candidate genes directly in mapped QTLs regions.
A total of 1733 F1 seedlings derived from 'Jonathan' × 'Golden Delicious' was used for the map constructionand QTL analysis. The SNP markers were developed by restriction site-associated DNA sequencing (RADseq). Phenotyping data of fruit quality traits were calculated in 2008-2011. Once QTLs were mapped, candidate genes were searched for in the corresponding regions of the apple genome sequence underlying the QTLs. Then some of the candidate genes were validated using real-time PCR.
A high-density genetic map with 3441 SNP markers from 297 individuals was generated. Of the 3441 markers, 2017 were mapped to 'Jonathan' with a length of 1343.4 cM and the average distance between markers was 0.67 cM, 1932 were mapped to 'Golden Delicious' with a length of 1516.0 cM and the average distance between markers was 0.78 cM. Twelve significant QTLs linked to the control of fruit weight, fruit firmness, sugar content and fruit acidity were mapped to seven linkage groups. Based on gene annotation, 80, 64 and 17 genes related to fruit weight, fruit firmness and fruit acidity, respectively, were analyzed.Among the 17 candidate genes associated with control of fruit acidity, changes in the expression of MDP0000582174 (MdMYB4) were in agreement with the pattern of changes in malic acid content in apple during ripening, and the relative expression of MDP0000239624 (MdME) was significantly correlated withfruit acidity.
We demonstrated the construction of a dense SNP genetic map in apple using next generation sequencing and that the increased resolution enabled the detection of narrow interval QTLs linked to the three fruit quality traits assessed. The candidate genes MDP0000582174 and MDP0000239624 were found to be related to fruit acidity regulation. We conclude that application of RADseq for genetic map construction improved the precision of QTL detection and should be utilized in future studies on the regulatory mechanisms of important fruit traits in apple.
基于遗传图谱的数量性状位点（QTL）分析是研究苹果重要园艺性状的重要方法。为了促进苹果果实品质性状的分子育种研究，我们旨在构建一张高密度图谱，该图谱对QTL定位有效，并且能够直接在定位的QTL区域搜索候选基因。
总共1733株由“乔纳金”ד金冠”杂交得到的F1实生苗用于图谱构建和QTL分析。通过限制性位点关联DNA测序（RADseq）开发SNP标记。2008 - 2011年计算果实品质性状的表型数据。一旦定位到QTL，就在QTL下方的苹果基因组序列相应区域搜索候选基因。然后使用实时PCR对一些候选基因进行验证。
构建了一张包含来自297个个体的3441个SNP标记的高密度遗传图谱。在这3441个标记中，2017个标记定位到“乔纳金”，长度为1343.4厘摩，标记间平均距离为0.67厘摩；1932个标记定位到“金冠”，长度为1516.0厘摩，标记间平均距离为0.78厘摩。12个与果实重量、果实硬度、糖含量和果实酸度控制相关的显著QTL定位到7个连锁群。基于基因注释，分别分析了80个、64个和17个与果实重量、果实硬度和果实酸度相关的基因。在与果实酸度控制相关的17个候选基因中，MDP0000582174（MdMYB4）的表达变化与苹果成熟过程中苹果酸含量的变化模式一致，MDP0000239624（MdME）的相对表达与果实酸度显著相关。
我们展示了利用下一代测序技术构建苹果高密度SNP遗传图谱，并且提高的分辨率能够检测到与所评估的三个果实品质性状相关的窄区间QTL。发现候选基因MDP0000582174和MDP0000239624与果实酸度调控有关。我们得出结论，RADseq应用于遗传图谱构建提高了QTL检测的精度，应在未来苹果重要果实性状调控机制的研究中加以利用。
