利用高温固相反应法制备出Ba3Bi2-x(PO4)4:xTb3+(x=0.05,0.1,0.15,0.3,0.4,0.5)绿色荧光粉.通过X射线衍射仪、扫描电子显微镜、积分球式分光光度计和荧光光谱仪等对样品进行了分析.结果表明,所制备的样品均为Ba3Bi2(PO4)4纯相,Ba3Bi1.7(PO4)4:0.3Tb3+的带隙估计值为4.21 eV.当激发光的波长为377 nm时,样品的发射光谱的波峰位于543 nm、584 nm和619 nm处,分别对应于Tb3+的5 D4→7 F5、5 D4→7 F4和5 D4→7 F3的能级跃迁.随着Tb3+掺杂浓度的增加,样品的发光强度先增强后减弱,当x=0.3时,发光强度最大.计算表明最近邻离子在Ba3 Bi2-x(PO4)4:xTb3+荧光粉的浓度猝灭中起主要作用.随着测试温度的升高,发光强度变化不大,表明样品具有优异的热稳定性能.CIE色坐标图表明所制备的样品可以被紫外光有效激发而发出绿光.

[目的]探究烟草叶片数杂交优势表现,并分析烟草叶片数相关基因的差异表达情况及杂种优势形成的原因,为深入研究烟草叶片数的分子遗传基础和选育叶片数较多的杂交种提供理论依据.[方法]以叶片数差异较大的9个烟草品种(系)为亲本,按照NCⅡ遗传交配设计组配20个杂交组合,并测定亲本和杂交组合的叶片数,计算其杂种优势,从中筛选出强、弱优势组合,利用实时荧光定量PCR检测其叶片相关基因BRI1、BSK3、FLC、FPF1和PHYC的相对表达量.最后,对叶片数相关基因中亲表达优势间及其与叶片数中亲优势进行相关分析.[结果]9个亲本材料的叶片数为20.33～33.22片,以GDH94的叶片数最多,其次是南江三号和毕纳1号,三者间无显著差异(P＞0.05,下同),但GDH94显著高于其余6个亲本(P＜0.05,下同),表明供试亲本间的叶片数存在真实的遗传差异.20个杂交组合的叶片数存在明显差异,为20.89～31.33片,以GDH94×南江三号的叶片数最多,以NC82×青梗的叶片数最少,说明采用杂种优势育种方法可选育出烟草叶片数较多的杂交种.20个杂交组合叶片数的杂种优势差异较大,其中中亲优势为-14.71%～11.77%,表现为正向中亲优势和负向中亲优势的组合分别占25%和75%,其中,以K326×GDH88叶片数的正向中亲优势最强,为11.77%,以GDH94×湄潭大蛮烟叶片数的负向中亲优势最强,为-14.71%;NC82×南江三号叶片数的中亲优势最弱,为-0.22%,故选择K326×GDH88和GDH94×湄潭大蛮烟为强优势组合、NC82×南江三号为弱优势组合.不同叶片数相关基因的中亲表达优势之间存在一定的相关性,其中BRI1和BSK3基因的中亲表达优势之间存在显著正相关;FPF1和PHYC基因的中亲表达优势与叶片数杂种优势存在显著负相关.FPF1基因在正向强优势杂交组合K326×GDH88和弱优势组合NC82×南江三号中较其相应亲本下调表达,但负向强优势组合GDH94×湄潭大蛮烟较其亲本上调表达. PHYC基因在正向强优势组合K326×GDH88和弱优势组合NC82×南江三号中较其相应亲本下调表达,但在负向强优势组合GDH94×湄潭大蛮烟较其亲本上调表达.[结论]K326×GDH88组合的叶片数杂种优势最大,具有较大的高产潜力.FPF1和PHYC基因参与调控烟草叶片数杂种优势的形成,其下调表达是烟草叶片数性状杂种优势形成的分子基础,可指导亲本选配,提高烟草杂交选育效率.

烟草是一种对光照和温度要求都非常严格的喜光喜温作物,为了降低烟草品种对光照和温度的敏感性,扩大烟草品种的光温适应范围和种植区域.本研究克隆了 NtPHYA基因,采用生物信息学方法预测了该基因在烟草上的功能,利用CRISPR/Cas9基因编辑技术创制NtPHYA基因敲除突变体,分析了 NtPHYA基因对烟草种子和烟苗光温敏感性及烟株生长的影响.结果表明,烟草NtPHYA与番茄SlPHYA基因的蛋白质序列相似度达92.15％,具有极为相似的理化性质,推测NtPHYA基因与番茄PHYA基因一样具有调控光温度敏感性和生长发育的功能;NtPHYA基因敲除突变体的光合色素含量、光合特性指标以及主要农艺性状均显著高于野生型,不同光温处理下突变体的NtPHYA基因表达量、种子萌发率均显著高于野生型.结果说明NtPHYA基因具有调控烟草光温度敏感性的功能,敲除NtPHYA基因能增加烟草光合色素含量、提高光合效率、促进烟株的生长,拓宽了烟草的光温适应范围和种植区域.本研究结果为解析NtPHYA基因对烟草生长发育的调控机理提供了理论参考和研究材料,对光温低敏感性烟草品种的选育具有重要的实践意义.

The sterile line is the basis of crop heterosis utilization. To broaden the sources of male sterility in tobacco, the Ntms1 (Nicotiana tabacum L. ms1) gene was cloned from the tobacco variety K326 by homologous cloning based on the Cams1 ( Capsicum annuum L. ms1 ) gene sequence of male-sterility genes in pepper. The protein structure and physicochemical properties of the two genes were determined by bioinformatics analysis, and the function of the Ntms1 gene was verified by the CRISPR/Cas9 system. The results showed that the sequences of Ntms1 and Cams1 were 85.25% similar, and plant homeodomains were found in both genes; the physical and chemical properties were also very similar. It is speculated that the Ntms1 gene had the same function as the Cams1 gene in controlling male sterility. Compared to the wild-type plants, the filaments of the Ntms1 knockout mutant plants were shorter, and the stamen was shorter than the pistil. The anthers did not develop fully and had few viable pollen grains; the tapetum and the anther wall had developed abnormally, and the anther chamber was severely squeezed. The malondialdehyde content in the mutant plants was significantly higher than that in the wild-type plants, while self-fertility was significantly lower in the mutant plants. The results showed that the Ntms1 gene plays an important role in regulating fertility in tobacco.

杂种优势利用是提高作物产量和品质的一种重要育种方法,明确生态条件和打顶措施对杂种优势的影响,对提高烟草杂种优势研究利用效率有重要意义.本试验以烟叶烟碱含量杂种优势相差较大的3个杂交组合及其亲本为材料,选择生态条件差异较大的3个生态试验点,研究了不同生态环境和打顶措施对烟叶烟碱含量、烟叶烟碱含量杂种优势及其相关基因相对表达量的影响.结果表明,不同生态环境间的烟叶烟碱含量差异显著,烟株打顶前后烟叶烟碱含量发生了显著的变化,打顶处理的烟叶烟碱含量显著高于不打顶处理;不同生态环境、打顶和不打顶处理间的烟叶烟碱含量的中亲优势值差异不显著,而超亲优势值、超标优势值差异显著;不同生态环境、打顶和不打顶处理间,烟叶烟碱含量杂种优势相关的PM T、A O、QS基因的中亲优势表达量相差也不显著,与处理间中亲优势值的表现差异一致.结果表明,在选用中亲优势作为度量指标进行杂种优势的遗传研究中,可不考虑生态环境和打顶的影响,在一个试验点对双亲及F1采用相同的农艺措施就能获得较为准确的鉴定结果;而在选用超亲优势、超标优势作为度量指标进行杂交种应用潜力评价中,需在不同生态环境、不同农艺措施下进行鉴定.

Heterosis is a common biological phenomenon that can be used to optimize yield and quality of crops. Using heterosis breeding, hybrids with suitable nicotine content have been applied to tobacco leaf production. However, the molecular mechanism of the formation of nicotine heterosis has never been explained from the perspective of protein. The DIA proteomics technique was used to compare the differential proteomics of the hybrid Va116 × Basma, showing strong heterosis in nicotine content from its parent lines Va116 and Basma. Proteomics analysis indicated that 65.2% of DEPs showed over-dominant expression patterns, and these DEPs included QS, BBL, GS, ARAF and RFC1 which related to nicotine synthesis. In addition, some DEPs (including GST, ABCE2 and ABCF1 and SLY1) that may be associated with nicotinic transport exhibited significant heterosis over the parental lines. These findings demonstrated that the efficiency of the synthesis and transport of nicotine in hybrids was significantly higher than that in the parent lines, and the accumulation of over-dominant expression proteins may be the cause of heterosis of nicotinic content in hybrids.

In order to explore the molecular genetic basis of proline heterosis in tobacco under drought stress, 7 tobacco varieties with different drought resistance and 10 hybrid combinations were used as materials, and drought stress treatments were set up to analyze proline heterosis and differential expression of related genes in tobacco by pot experiments. The results showed that the content of tobacco proline under drought stress increased continuously at the early stage and decreased after the 14th day. The heterosis of proline content in hybrid combinations was significantly different, and the proline content had the mid-parent heterosis with the highest value of 40.35 and the lowest of -51.66. The relative expression levels of the proline synthesis key enzyme genes P5CS and δ-OAT were significantly higher in the strong-dominant combinations, being 2.10 times and 1.87 times of those of the weak-dominant combinations,respectively.There was a significant positive correlation between the proline heterosis and P5CS gene relative expression after persistent drought for 7d at the vigorous growth stage. The correlations between the proline heterosis and δ-OAT gene relative expression were significant or extremely significant after persistent drought for 14d at the root extension stage and persistent drought for 7d and 14d at the vigorous growth, with correlation coefficients of 0.93, 0.98 and -0.96, respectively. Therefore, the up-regulation or down-regulation of δ-OAT and P5CS was the molecular basis for the formation of heterosis of tobacco proline content during the corresponding growth periods.