Mapping and transcriptomic profiling reveal that the KNAT6 gene is involved in the dark green peel colour of mature pumpkin fruit (Cucurbita maxima L.)

IF 4.4 1区 农林科学 Q1 AGRONOMY Theoretical and Applied Genetics Pub Date : 2024-09-17 DOI:10.1007/s00122-024-04741-7
ChaoJie Wang, Wenqi Ding, Fangyuan Chen, Ke Zhang, Yuetong Hou, Guichao Wang, Wenlong Xu, Yunli Wang, Shuping Qu
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Key message

We identified a 580 bp deletion of CmaKNAT6 coding region influences peel colour of mature Cucurbita maxima fruit.

Abstract

Peel colour is an important agronomic characteristic affecting commodity quality in Cucurbit plants. Genetic mapping of fruit peel colour promotes molecular breeding and provides an important basis for understanding the regulatory mechanism in Cucurbit plants. In the present study, the Cucurbita maxima inbred line ‘9-6’ which has a grey peel colour and ‘U3-3-44’ which has a dark green peel colour in the mature fruit stage, were used as plant materials. At 5–40 days after pollination (DAP), the contents of chlorophyll a, chlorophyll b, total chlorophyll and carotenoids in the ‘U3-3-44’ peels were significantly greater than those in the ‘9-6’ peels. In the epicarp of the ‘9-6’ mature fruit, the presence of nonpigmented cell layers and few chloroplasts in each cell in the pigmented layers were observed. Six generations derived by crossing ‘9-6’ and ‘U3-3-44’ were constructed, and the dark green peel was found to be controlled by a single dominant locus, which was named CmaMg (mature green peel). Through bulked-segregant analysis sequencing (BSA-seq) and insertion-deletion (InDel) markers, CmaMg was mapped to a region of approximately 449.51 kb on chromosome 11 using 177 F2 individuals. Additionally, 1703 F2 plants were used for fine mapping to compress the candidate interval to a region of 32.34 kb. Five coding genes were in this region, and CmaCh11G000900 was identified as a promising candidate gene according to the reported function, sequence alignment, and expression analyses. CmaCh11G000900 (CmaKNAT6) encodes the homeobox protein knotted-1-like 6 and contains 4 conserved domains. CmaKNAT6 of ‘9-6’ had a 580 bp deletion, leading to premature transcriptional termination. The expression of CmaKNAT6 tended to increase sharply during the early fruit development stage but decrease gradually during the late period of fruit development. Allelic diversity analysis of pumpkin germplasm resources indicated that the 580 bp deletion in the of CmaKNAT6 coding region was associated with peel colour. Subcellular localization analysis indicated that CmaKNAT6 is a nuclear protein. Transcriptomic analysis of the inbred lines ‘9-6’ and ‘U3-3-44’ indicated that genes involved in chlorophyll biosynthesis were more enriched in ‘U3-3-44’ than in ‘9-6’. Additionally, the expression of transcription factor genes that positively regulate chlorophyll synthesis and light signal transduction pathways was upregulated in ‘U3-3-44’. These results lay a foundation for further studies on the genetic mechanism underlying peel colour and for optimizing peel colour-based breeding strategies for C. maxima.

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图谱和转录组分析表明,KNAT6 基因与成熟南瓜果实(Cucurbita maxima L.)深绿色果皮的颜色有关
摘要果皮颜色是影响葫芦科植物商品质量的重要农艺性状。果皮颜色的遗传图谱促进了分子育种,并为了解葫芦科植物的调控机制提供了重要依据。本研究以果皮颜色为灰色的 Cucurbita maxima 近交系'9-6'和成熟果实阶段果皮颜色为深绿色的'U3-3-44'为植物材料。授粉后 5-40 天,'U3-3-44'果皮中叶绿素 a、叶绿素 b、总叶绿素和类胡萝卜素的含量明显高于'9-6'果皮。在'9-6'成熟果实的外果皮中,观察到存在非色素细胞层,色素层中每个细胞中的叶绿体数量很少。通过构建'9-6'和'U3-3-44'杂交产生的六代,发现深绿色果皮由一个显性基因座控制,该基因座被命名为 CmaMg(成熟绿色果皮)。通过大量分离分析测序(BSA-seg)和插入-缺失(InDel)标记,利用177个F2个体将CmaMg映射到11号染色体上约449.51 kb的区域。此外,还利用 1703 株 F2 进行了精细图谱绘制,将候选区间压缩到 32.34 kb 的区域。该区域有五个编码基因,根据报告的功能、序列比对和表达分析,CmaCh11G000900 被确定为有希望的候选基因。CmaCh11G000900 (CmaKNAT6)编码同源框蛋白结-1-样 6,包含 4 个保守结构域。9-6'的 CmaKNAT6 有 580 bp 的缺失,导致转录过早终止。CmaKNAT6 的表达量在果实发育早期急剧增加,但在果实发育后期逐渐减少。南瓜种质资源的等位基因多样性分析表明,CmaKNAT6编码区的580 bp缺失与果皮颜色有关。亚细胞定位分析表明,CmaKNAT6 是一种核蛋白。对近交系 "9-6 "和 "U3-3-44 "的转录组分析表明,与 "9-6 "相比,"U3-3-44 "中参与叶绿素生物合成的基因更为丰富。此外,'U3-3-44'中对叶绿素合成和光信号转导途径有正向调节作用的转录因子基因的表达上调。这些结果为进一步研究果皮颜色的遗传机制和优化基于果皮颜色的 C. maxima 育种策略奠定了基础。
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来源期刊
CiteScore
9.60
自引率
7.40%
发文量
241
审稿时长
2.3 months
期刊介绍: Theoretical and Applied Genetics publishes original research and review articles in all key areas of modern plant genetics, plant genomics and plant biotechnology. All work needs to have a clear genetic component and significant impact on plant breeding. Theoretical considerations are only accepted in combination with new experimental data and/or if they indicate a relevant application in plant genetics or breeding. Emphasizing the practical, the journal focuses on research into leading crop plants and articles presenting innovative approaches.
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