人类对食品的“色-香-味”有着强烈而执著的追求，因此改善作物的“色-香-味”性状一直是育种家梦寐以求的目标。然而长期以来育种研究多局限于“色泽育种”、“香味育种”及“甜味育种”等单一性状的优化，难以满足消费者对食物色香味的整体需求。本研究采用分子生物学和转基因技术，首次构建了包含“色-香-味”三个性状的4基因共表达系统，培育出同时改善番茄“色-香-味”三性状的转基因株系，为作物多性状遗传改良提供了新的研究思路。
“色”是作物最直观的性状，选择合适的显色成分是研制“色-香-味”三性状转基因株系的基础和关键，同时植物色素也是值得单独开发利用的天然食品添加剂。在众多植物色素中，类胡萝卜素类色素西红花苷以及黄酮类色素黄芩素等因具有较好的药理活性而被广泛关注。西红花苷不仅具有抗抑郁和治疗阿尔茨海默病的潜力，还展现出较好的抗氧化和抗炎等多种生物活性；黄芩素则具有抗菌、抗病毒、预防糖尿病及治疗其并发症等功效。通过在植物底盘异源合成药用色素，既能解决其资源短缺问题，也可赋予食品保健功能。
本研究以西红花苷、黄芩素和花青素为起点，探索前两者异源生物合成的潜力，同时评价三种色素的显色效果。在评估了不同色素的显色效应之后，我们选择显色更显著且具抗氧化活性的花青素作色泽模块。香味模块选用玫瑰精油主成分香叶醇，其兼具花香特性、抗菌抗癌活性及提升植物抗病性的潜力；甜味模块则引入零卡高甜度的甜蛋白Thaumatin II。上述三种“色-香-味”成分均具有一定的生理功效，其基因可为功能型蔬菜的选育提供基因供体。
本文通过多基因载体构建及遗传转化技术探讨了西红花苷和黄芩素的异源生物合成，以及“色-香-味”三成分在番茄中的协同合成，主要研究结果如下：
（1）在烟草/番茄中实现高比例西红花苷I和II的异源生物合成
将栀子中西红花苷合成的4个关键基因GjCCD4a、GjALDH2C3、GjUGT74F8和GjUGT94E13组装到一个多基因植物表达载体上，并通过基因工程技术转入番茄和烟草中；经抗性筛选及分子鉴定，获得5株阳性烟草植株（N16、N18、N20、N22、N34）和7株阳性番茄植株（S5、S7、S10、S13、S15、S24、S25）。烟草株系N16与N18主要积累西红花苷I，其T0、T1代植株中的西红花苷I含量占总苷的83%-89%；株系N20与N22以西红花苷II为主，含量占总苷的66%和77%。T1代烟草株系N16-HT1中西红花苷的鲜重为78.36 μg/g（fresh weight, FW），其中西红花苷I和II占总苷含量的99%。番茄株系S10果实中西红花苷含量为37.13 μg /g FW，其中西红花苷I占总苷含量的98%（株系S15达99%）。
（2）首次在番茄中实现黄芩素的异源生物合成
将黄芩素生物合成通路的关键基因（SbCLL-7、SbCHI、SbCHS-2、SbFNSII-2、SbCYP82D1.1）整合至pCAMBIA1300载体上，获得多基因植物表达载体FCC-CF，通过农杆菌介导的遗传转化技术将FCC-CF转入番茄；经抗性筛选与分子鉴定，共获得3株阳性转基因株系（F8、F12、F21）。HPLC-MS/MS分析表明，转基因番茄果实中黄芩素含量最高达558 ng/g FW。
（3）首次建立了改善番茄“色-香-味”三性状的转基因株系
在比较了三种色素的显色效果后，确定采用花青素作为色素供体。本研究通过将花青素调控基因 SlMYB75（显色）、香叶基二磷酸合成酶基因AgGPPS2（前体供应）与 ObGES（香气生成）以及甜味蛋白基因 thaumatin II（增甜），构建为多基因载体GT-MG，并利用遗传转化技术获得4株转基因番茄株系（G6、G12、G28、G35）。PCR鉴定及定量逆转录PCR（quantitative reverse transcription PCR, qRT-PCR）分析证实“色-香-味”性状基因均成功整合到番茄基因组中并成功表达。植株根、茎、叶及果实均呈现明显的紫色性状，花青素含量较野生型显著提升；GC-MS分析显示，转基因株系叶片中香叶醇含量高达1215.14 ng/g FW，果实发育阶段中，绿熟期、破色期及破色期后8-10天的香叶醇含量分别为野生型同期果实的271倍、16倍和13倍；酶联免疫吸附测定（enzyme-linked immunosorbent assay, ELISA）证实，所有转基因株系果实中均成功合成甜蛋白 Thaumatin II；实现了同时改善“色-香-味”三性状的技术创新。
综上，本研究获得了高比例西红花苷I和II的转基因番茄/烟草株系，为后续定向开发高纯度产品奠定基础，转基因番茄果实呈现“红中带金”的独特色泽，也具有开发为彩色功能性食品的潜力。本研究首次实现了黄芩素在植物底盘中的异源生物合成，为含黄芩素功能食品的开发奠定了基础。更重要的是实现了花青素、香叶醇和甜蛋白Thaumatin II在番茄中的协同合成，成功构建了“色-香-味”组合性状的转基因番茄株系，为风味育种和功能性食品开发提供了新的路径和经验。

Humans have a strong and persistent pursuit of the "color-aroma-taste" of food. Consequently, improving these sensory traits in edible plants has long been a desirable goal in plant breeding. However, traditional breeding efforts have mostly focused on the improvement of single traits—color, aroma, or sweetness individually, which falls short of meeting consumer demand for comprehensive flavor quality. In this dissertation, we employed molecular biology and genetic engineering approaches to construct, for the first time, a multi-gene co-expression system integrating four genes responsible for "color-aroma-taste" traits. We successfully developed transgenic tomato lines with altered "color-aroma-taste" characteristics, offering a new strategy for multi-trait genetic improvement in crops. 
Among these traits, "color" is the most visually perceivable trait of crops. Selecting appropriate pigment components is fundamental and essential for developing transgenic lines with integrated "color-aroma-taste" traits. Meanwhile, plant pigments also represent natural food additives worthy of independent development and utilization. Among various plant pigments, carotenoid pigments such as crocins and flavonoid pigments like baicalein have attracted widespread attention due to their significant pharmacological activities. Crocins possess a range of biological activities, including antidepressant effects, potential for treating Alzheimer’s disease, and strong antioxidant and anti-inflammatory properties. Baicalein exhibits antibacterial, antiviral, antidiabetic, and treatment of diabetic complications effects. The heterologous synthesis of medicinal pigments in plant chassis can address resource limitations while also endowing food with health protective effects.
This research targeted crocin, baicalein, and anthocyanins, exploring the feasibility of heterologous biosynthesis for the first two pigments while comparatively evaluating the coloration characteristics of all three. Following assessment of coloration effects across different pigments, we selected anthocyanins which exhibit visually distinctive coloration and possess antioxidant activity as the color module. Geraniol, the primary component of rose essential oil, was selected as the aroma module due to its floral fragrance, anti-inflammatory and antibacterial properties, and potential to enhance disease resistance in plants. The zero-calorie sweet protein Thaumatin II was employed as the taste module. These components each exhibit specific physiological activities, so their corresponding genes can serve as donors for the breeding of functional vegetables.
This research investigates the heterologous biosynthesis of crocins and baicalein via multigene vector construction and genetic transformation techniques, and explores the coordinated synthesis of "color-aroma-taste" components in tomato. The main findings are as follows:
(1) Achieve heterologous biosynthesis of crocins I and II with high proportions in tobacco/tomato.
Four key genes involved in crocins biosynthesis in Gardenia jasminoides—GjCCD4a, GjALDH2C3, GjUGT74F8, and GjUGT94E13—were assembled into a multi-gene plant expression vector. This construct was introduced into tobacco and tomato plants via genetic engineering. Following antibiotic selection and molecular verification, five positive tobacco lines (N16, N18, N20, N22, N34) and seven positive tomato lines (S5, S7, S10, S13, S15, S24, S25) were obtained. Tobacco lines N16 and N18 primarily accumulated crocin I, with crocin I content constituting 83%-89% of total crocins in both T0 and T1 generations. Lines N20 and N22 predominantly produced crocin II, accounting for 66% and 77% of total crocins, respectively. In the T1 tobacco line N16-HT1, total crocins content reached 78.36 μg/g (FW), with the combined content of crocins I and II accounting for 99%. In tomato line S10, content of crocins in the fruit reached 37.13 μg/g FW, with crocin I accounting for 98% (99% in line S15).
(2) First achievement of heterologous biosynthesis of baicalein in tomatoes.
Five key genes from the baicalein biosynthetic pathway (SbCLL-7, SbCHI, SbCHS-2, SbFNSII-2, SbCYP82D1.1) were assembled into the pCAMBIA1300 vector to construct the multi-gene expression cassette FCC-CF. Using Agrobacterium-mediated transformation, FCC-CF was introduced into tomato. After selection and molecular analysis, three positive transgenic lines (F8, F12, F21) were identified. HPLC-MS/MS analysis revealed that baicalein content in the transgenic tomato fruit reached up to 558 ng/g FW.
(3) First construction of transgenic tomato lines with "color-aroma-taste" traits.
After comparing the coloration effects of the three pigments, anthocyanin was selected as the pigment donor. This study achieved technical innovation by constructing a multigene vector GT-MG containing anthocyanin regulatory gene SlMYB75 (color enhancement), geranyl diphosphate synthase AgGPPS2 (precursor supply) and ObGES (aroma generation), along with sweet-tasting protein gene thaumatin II (sweetness improvement). Four transgenic tomato lines (G6, G12, G28, G35) were obtained through Agrobacterium-mediated transformation.
PCR identification and qRT-PCR (quantitative reverse transcription PCR) analysis confirmed the successful integration and expression of "color-aroma-taste" trait genes in the tomato genome. These transgenic lines exhibited pronounced purple coloration in roots, stems, leaves, and fruits, with significantly elevated anthocyanin levels compared to wild-type plants. GC-MS analysis revealed that the geraniol content in the leaves of the transgenic line reached as high as 1215.14 ng/g FW, which was 558 times that of the wild-type. In fruits, the geraniol content peaked at 271 times, 16 times, and 13 times that of the wild-type during the green mature stage, breaker stage, and 8-10 days after the breaker stage, respectively. enzyme-linked immunosorbent assay (ELISA) results confirmed successful synthesis of Thaumatin II in all fruit samples. This study achieved a technological breakthrough in simultaneous improvement of three key quality traits: coloration, aroma, and sweetness.
In summary, this research successfully obtained transgenic tobacco/tomato lines with high proportions of crocin I and II, laying the foundation for subsequent targeted development of high-purity products. The transgenic tomato fruit exhibited a unique "gold-tinted red" coloration, demonstrating potential for development as chromatic functional foods. Notably, this research achieved the first successful heterologous biosynthesis of baicalein in a plant chassis system, establishing a crucial foundation for functional food production containing this bioactive compound. More importantly, we realized the coordinated synthesis of anthocyanins, geraniol, and sweet-tasting protein Thaumatin II in tomato, creating transgenic lines with combined "color-aroma-taste" and offering a new direction and experience for flavor breeding and the production of functional foods.