甘蓝型油菜Cu/ZnSOD和FeSOD基因的克隆及菌核病菌诱导表达

依据拟南芥、芥菜型油菜和白菜已知超氧化物歧化酶(SOD)保守序列设计引物,用同源序列法和RT-RACE技术克隆甘蓝型油菜Cu/ZnSOD和FeSOD基因,经序列分析和基因片段拼接,得到Cu/ZnSOD和FeSOD基因的全长cDNA,分别为756 bp (GenBank登录号AY970822)和1 037 bp (GenBank登录号EF634058)。以cDNA序列设计引物,获得1 322 bp的Cu/ZnSOD基因组DNA (GenBank登录号DQ431853)和1 659 bp的FeSOD基因组DNA (GenBank登录号EF634057)。生物信息学分析表明,Cu/ZnSOD基因ORF框长459 bp,编码152个氨基酸残基的蛋白质,在基因组序列结构上具有7个外显子和6个内含子。而FeSOD基因ORF框长792 bp,编码263个氨基酸残基的蛋白质,在基因组序列结构上具有8个外显子和7个内含子。二者外显子和内含子交接处完全符合GT/AG规则。利用获得的Cu/ZnSOD的cDNA片段作探针,对菌核病菌诱导甘蓝型油菜叶片的mRNA进行Northern blotting分析,结果显示在同一品种(系)中菌核病菌诱导后Cu/ZnSOD mRNA表达量比诱导前升高,抗(耐)型油菜Cu/ZnSOD mRNA表达量明显高于感病型。油菜叶片SOD酶活性分析结果也获得了完全一致的结果。以上结果表明,甘蓝型油菜SOD基因与菌核病抗性相关。     

油菜转录因子BnZFP基因的分离及其功能初步分析

为克隆油菜ZFP类转录因子基因,初步阐明其功能。采用RT-PCR的方法克隆基因和体内检测;花沾法转化拟南芥,Northern blotting检测转基因后的表达。从甘蓝型油菜中分离了一个新的ZFP类转录因子,它编码377个氨基酸,与拟南芥AtZFP基因(At2g29660)的碱基序列相似性最高,为76.2%,命名为BnZFP。该基因在父本、母本和F1代不同发育时期中均能较稳定的表达;该基因转化拟南芥,对获得的转BnZFP基因拟南芥进行了Northern Blotting分析,表明该基因在拟南芥中得到过表达;与野生型拟南芥相比较,过表达BnZFP的转基因拟南芥在萌发、营养生长及生殖生长过程中没有表现出明显的表型变化,该基因的功能有待进一步研究。     

甘蓝型油菜甜菜碱醛脱氢酶基因(BnBADH-1)的克隆与真核表达载体的构建

根据已知的拟南芥甜菜碱醛脱氢酶基因(BADH1)序列设计特异性引物,利用RT-PCR技术从甘蓝型油菜中克隆出油菜甜菜碱醛脱氢酶基因(BnBADH-1)全长。测序和分析结果表明,BnBADH-1的cDNA全长1506bp,编码一个包含501个氨基酸残基、分子量为55kD、等电点(pI)为5.16的假定蛋白质;核酸序列和氨基酸序列与拟南芥的同源性分别为90.24%和93.41%。利用片段两端的粘性末端酶切位点BamHⅠ和SacⅠ将cDNA片段连接在真核表达载体pBI121上;PCR鉴定表明,过量表达载体pBI121-BnBADH-1已成功构建,为下一步的基因功能分析做好了准备。     

棉纤维发育相关基因GhCHS、GhCPI的克隆与鉴定

  以陆地棉李氏超短纤维突变体Li₁li₁自交后代野生型li₁li₁和超短纤维突变体Li₁li₁开花后4 d的胚珠纤维的复合体的RNA为探针,通过基因芯片的方法筛选优质材料7235棉纤维伸长、次生壁加厚不同发育时期混合cDNA文库,分离出两个在两种材料中差异表达的cDNA序列,分别命名为GhCHS（GenBank登录号：EF643506）和GhCPI（GenBank登录号：EF643507）。RT-PCR分析表明：开花后4 d,GhCHS,GhCPI在陆地棉李氏野生型li₁li₁纤维中的表达量低于超短纤维突变体Li₁li₁。Southern杂交结果表明两个基因在陆地棉基因组中都存在两个拷贝     

甘蓝型油菜BnaABI4基因CRISPR/Cas9编辑载体的构建及甘蓝型油菜转化

ABI4(Abscisicacid-insensitive4)是ABA响应途径中重要的转录因子。本研究以甘蓝型油菜全基因组数据库为基础,借助拟南芥ABI4蛋白的氨基酸序列,通过序列比对,获得了甘蓝型油菜的2个BnaABI4,分别命名为BnaABI4-A (GenBank登录号为BnaA03g18970D)和BnaABI4-C (GenBank登录号为BnaC03g725-10D)。在这两个基因的CDS序列上寻找一段长度为20 bp、序列相同的片段作为基因编辑的目标位点,构建CRISPR/Cas9基因编辑载体pCAMBIA1300-sg RNA/Cas9-BnaABI4;借助根癌农杆菌介导的油菜下胚轴遗传转化法,成功将BnaABI4的基因编辑表达框转入甘蓝型油菜‘湘油15’中,共获得转基因植株13株,这将为深入研究甘蓝型油菜BnaABI4的生物学功能提供科学依据。     

Bna-novel-miR311-HSC70-1模块调控甘蓝型油菜响应热胁迫的机制

HSP70(heat shock protein 70)参与植物热胁迫应答,增强植物耐热性,但目前油菜中尚无miRNA调控HSP70基因的报道。本研究novel-miR311是利用高通量技术在甘蓝型油菜茎尖中筛选出的新miRNA。novel-miR311存在于油菜而不存在于拟南芥中, 5'-RACE技术证实其2个靶基因属热应激同源蛋白基因HSC70-1 (HSP70家族),在甘蓝型油菜体内被剪切。构建novel-miR311超表达载体,转化拟南芥和甘蓝型油菜,其转基因阳性苗中HSC70-1基因表达量显著下降。高温胁迫试验表明,拟南芥和甘蓝型油菜热胁迫后,其阳性苗的生长势和存活率显著低于其对应的对照。qPCR结果显示,油菜中HSC70-1基因表达量热胁迫后较热胁迫前上升。上述结果表明,油菜novel-miR311介导HSC70-1基因的剪切降低了拟南芥和甘蓝型油菜耐热性。     

金鱼草AmPIN1a基因过表达载体的构建及转基因分析

本研究以金鱼草的AmPIN1a为目的基因,AmPIN1a基因全长CDS为1 836 bp,通过热激法(电击发)将构建过表达载体pCAMBIA1302 (1301)-AmPIN1a导入根癌农杆菌GV3101中,采用叶盘法转化拟南芥,观察其表型并用外源IAA对其进行处理。结果表明,转化AmPIN1a基因的拟南芥突变体在0～12 d内主根长度略短于野生型主根长度,而且侧根发育明显增多。移栽后,AmPIN1a拟南芥突变体的前期叶片大小也小于野生型,24 d后叶片逐步超过野生型,花序轴萌发晚,但发育较快。过表达AmPIN1a基因的拟南芥在外缘IAA处理下表现出比野生型拟南芥更敏感的性状,由此推测AmPIN1a基因可能在生长素运输过程中起作用。     

甘蓝型油菜BnADH3基因的克隆及转BnADH3拟南芥的耐淹性

利用RT-PCR技术从甘蓝型油菜耐淹品系WR-4中克隆获得Bn ADH3基因,其完整开放阅读框为1137 bp。该基因编码379个氨基酸,与甘蓝Bo ADH3基因和拟南芥At ADH3基因高度同源,同源性分别达到96%和91%。利用定量RT-PCR检测Bn ADH3基因在油菜耐淹系WR-4和不耐淹系WR-24中的表达表明,在淹水处理下该基因表达受到一定的诱导,淹水处理6 h后表达开始上调,说明Bn ADH3基因在油菜耐淹机制中发挥作用;将其转化到模式生物拟南芥中,采用幼苗淹水3 d后去水处理,测定表明转基因株系叶片和根系中乙醇脱氢酶活性均高于野生型;生长4周和6周的拟南芥植株淹水3 d后的表型显示,Bn ADH3的表达可增强拟南芥对淹水胁迫的耐性,处理的T2代转基因幼苗大部分恢复,但野生型幼苗枯死;调查表明,淹水5 d后野生型植株的存活率为26.7%,转基因株系ADH33和ADH44的存活率分别为80.0%和66.7%。     

甘蓝型油菜BnaFIL基因启动子的克隆与表达分析

为了进一步了解启动子在甘蓝型油菜FIL基因(BnaFIL)表达调控中的作用,根据甘蓝型油菜基因组数据,以甘蓝型油菜叶片提取的DNA为模板,对甘蓝型油菜BnaFIL基因的启动子序列pBnaFIL进行克隆,长度为1 326 bp。采用PlantCARE在线分析软件对该启动子序列进行生物信息学序列分析,结果表明,该序列含有参与光反应的部分保守DNA模块以及CAAT-box和TATA-box等核心启动子必备元件,与分生组织表达有关的顺式作用的调控元件CAT-box以及光敏反应元件。通过该启动子序列替换pBI121植物表达载体上的CaMV35S启动子,使该启动子与GUS基因融合获得pBnaFIL-GUS表达载体,将载体通过农杆菌花序浸染的方法转入拟南芥中,获得了早花启动子重组质粒阳性转基因株系和晚花启动子重组质粒阳性转基因株系。之后对转基因拟南芥植株进行GUS染色分析,对启动子的表达效果进行了检测,最终在不同的转基因拟南芥植株中均发现了GUS基因的表达。结果表明,早花材料与晚花材料中启动子表达强弱存在差异,早花材料启动子的驱动基因表达效果比晚花材料启动子的驱动效果要好,由此推断,启动子的驱动效果...     

甜菜亚硝酸还原酶基因(NiR)的克隆与表达分析

以甜菜品种甜研7号叶片的cDNA为模板,采用RT-PCR和3′/5′RACE技术,获得了编码亚硝酸还原酶基因(NiR)的cDNA全序列2 014 bp, 包含有1 830 bp的开放阅读框,编码599个氨基酸。所推导的氨基酸序列与菠菜及拟南芥NiR编码的氨基酸序列均具93%的同源性。生物信息学分析表明,甜菜NiR具有完整的NiR蛋白结构,含血红素蛋白β-化合物区域和4Fe-4S区域, 并利用分析软件预测其三维结构。实时荧光定量结果显示,在以0、10、20、30、40、50、80和160 mmol L^–1 NO₃^–-N处理72 h的试验中,50 mmol L^–1处理可使甜菜NiR的表达量达到最大;以0、2、4、8、16、32、64和128 mmol L^–1 NH₄⁺-N处理48 h的试验表明,8 mmol L^–1和64 mmol L^–1处理条件下甜菜NiR表达量相对较高。硝态氮和铵态氮不同配比处理48 h的试验中,NO₃^–-N和NH₄⁺-N比例为80:20可使甜菜NiR的表达量达到最大;在氮素诱导的基础上,蛋白抑制剂放线菌酮处理9 h,随着处理浓度的增大,NiR的表达量逐渐下降;不同浓度NO₂^–处理的试验中,40 mmol L^–1处理下NiR的表达量最大。     

西藏昌都市类乌齐牦牛肉中主要脂肪酸的差异分析

探讨西藏昌都市类乌齐县不同乡镇和不同部位牦牛肉中脂肪酸的差异,明确类乌齐牦牛肉脂肪酸的特征。采用气相色谱串联质谱法测定类乌齐牦牛肉中脂肪酸的含量,用SPSS方法分析不同乡镇和不同部位牦牛肉脂肪酸的差异。结果表明：类乌齐牦牛肉主要含有11种饱和脂肪酸(SFA)、7种单不饱和脂肪酸(MUFA)和9种多不饱和脂肪酸(PUFA),平均含量分别为42.53%、53.27%和4.21%。类乌齐牦牛肉中脂肪酸的主要组成为油酸(C18:1n9c,44.47%)、硬脂酸(C18:0,21.08%)和棕榈酸(C16:0,14.78%)。饱和脂肪酸和单不饱和脂肪酸总量在各乡镇之间差异不显著,多不饱和脂肪酸总量在各乡镇之间存在显著差异。多不饱和脂肪酸主要为亚油酸(2.74%)、花生四烯酸(0.66%)、亚麻酸(0.39%)、二十碳五烯酸(0.11%)和二十二碳六烯酸(0.07%)等。类乌齐牦牛肉中n-6/n-3均值为4.85,其臀部肉和腱子肉的n-6/n-3均值分别为3.20和3.35。类乌齐牦牛肉中油酸和硬脂酸含量较高,不同乡镇的牦牛肉中多不饱和脂肪酸差异显著,其臀部肉和腱子肉具有较高的营养价值。     

一氧化氮和亚甲基蓝对盐胁迫抗虫棉根系的抗氧化能力的影响

 通过检测抗氧化酶活性和抗氧化物含量,研究一氧化氮和亚甲基蓝对盐胁迫抗虫棉根系的抗氧化能力的影响。在正常条件和盐胁迫条件下分别用一氧化氮和亚甲基蓝处理抗虫棉幼苗,检测抗坏血酸(ASA)含量、抗坏血酸过氧化物酶(APX)活性、谷胱甘肽还原酶(GR)活性、谷胱甘肽(GSH/GSSG)值、脱氢抗坏血酸还原酶(DHAR)活性、单脱氢抗坏血酸还原酶(MDAR)活性,并作对比。结果表明：正常生长条件下添加NO能促进棉花幼苗生长,而添加亚甲基蓝(MB)显著抑制抗虫棉幼苗的生长;添加NO显著缓解了盐胁迫对棉花幼苗生长的抑制,提高了根系的抗氧化能力,盐胁迫下添加NO的同时添加MB可不同程度地降低以上抗氧化物质含量和抗氧化酶活力,解除NO对盐胁迫的促长作用。     

Wound-induced suberization genes are differentially expressed,spatially and temporally,during closing layer and wound periderm formation

Potato tuber (Solanum tuberosum L.) wounds incurred at harvest and upon seed cutting require rapid suberization as a major part of the healing process to prevent infection and desiccation. However, little is known about the induction and expression of genes that are essential for these processes and in particular to the two major stages of wound-induced suberization, i.e. closing layer formation and wound periderm formation. The objectives of this research were to address these needs by determining the effects of wounding on the induction and expression profiles of specific genes involved in wound-induced suberization in potato tuber (S. tuberosum L.) during the initiation and completion of closing layer formation and wound periderm formation. Although both stages critically involve suberization, there are significant differences between the two processes. Closing layer development requires rapid suberization of existing parenchyma cells bordering the wound surface to provide the initial protective barrier for the wound. Wound periderm development occurs later, i.e. after completion of closing layer formation, and requires development of a wound phellogen layer which mediates the formation of highly organized files of suberized wound-phellem cells that provide a more durable protective barrier for the tuber. The processes delineating these two separate stages of wound-induced suberization are poorly understood. This research shows that, unlike some wound responding genes such as phenylalanine ammonia lyase (StPAL-1) and anionic peroxidase (StPrx), certain genes that are specifically involved in both of these processes do not remain uniformly up-regulated during the two stages of healing (i.e. StTHT encoding Hydroxycinnamoyl-CoA:tyramine N-(hydroxycinnamoyl)transferase, StFHT encoding a fatty ω-hydroxyacid/fatty alcohol hydroxycinnamoyl transferase, StKCS6 encoding a 3-ketoacyl-CoA synthase, StFAOH encoding a fatty acid ω-hydroxylase and StGPAT5 encoding a protein with acyl-CoA:glycerol-3-phosphate acyltransferase). Instead, they are up-regulated during closing layer formation; i.e. starting by ca. 1 d after wounding, but then slightly down-regulated or pause near completion of the closing layer (ca. 5–6 d) and then again up-regulated as wound periderm development is fully initiated (ca. 7 d) and down-regulated near completion (ca. 28 d after wounding). This differential in the expression profile, i.e. decrease between stages, was not anticipated and may be the first demonstration of measurable changes of any sort of biological flux as wound induced suberization transitions from closing layer to wound periderm development. Results were repeated using minitubers from two different crop years and demonstrate that these processes are separate, but coupled in some yet to be determined fashion. The biology of this differential expression is important because of the roles closing layer and wound periderm development play in protecting the tuber from disease and other challenges.     

胡麻脂肪酸去饱和酶基因(fads)差异表达分析

为了了解fad基因在胡麻蒴果发育过程中对不饱和脂肪酸的调控,对高、中、低三个不同亚麻酸(C18:3)含量的胡麻品种(’CDC Gold’,‘内亚7号’,’Linola’)进行了不同时期的品质测定,以及脂肪酸去饱和酶2a基因(fad2a)、脂肪酸去饱和酶2b基因(fad2b)、脂肪酸去饱和酶2c基因(fad2c)、脂肪酸去饱和酶3a基因(fad3a)、脂肪酸去饱和酶3b基因(fad3b)的qRT-PCR定量分析。结果表明,随着蒴果成熟,可溶性糖含量呈降低趋势,粗脂肪与粗蛋白不断积累,且差异显著(p<0.05)。fad2a基因、fad3a基因以及fad3b基因在各个时期中的表达符合正态分布。以0 d的蒴果为对照,在胡麻种子形成过程中,‘内亚7号’的三个基因在5 d和15 d的表达量迅速增加,到30 d时急剧减少,15 d的fad2a基因表达量为5.23倍,fad3a基因表达量是fad2a基因表达量的14.52倍,fad3b基因的表达量是fad2a基因表达量的16.14倍,表明这三个基因参与不饱和脂肪酸积累过程。在低亚麻酸含量品种‘Linola’30 d中,fad3a基因是fad2a基因表达量的52.71倍,fad3b呈下调趋势;在高亚麻酸含量品种‘CDCGold’30d中,fad3a基因与fad2a基因的表达量均呈下调趋势,fad3b的表达量为3.92倍;在中等亚麻酸含量品种‘内亚7号’中,fad2a基因表达量降低了0.31倍,fad3a基因是fad3b基因表达量的1.87倍。fad2b基因、fad2c基因熔解曲线不稳定,峰值低,可以在后续试验中继续探索。     

不同生态气候区核桃坚果品质比较研究

为核桃跨生态气候区引种栽培提供技术参考,以产于亚热带生态气候区湖北保康县和暖温带生态气候区河北临城县早实核桃‘香玲’和晚实核桃‘清香’坚果为试材,采用连续两年的平均数据比较分析了它们的坚果表型数量性状、脂肪酸和氨基酸组分和含量及脂肪酸和氨基酸均衡性。结果表明：同一品种在两个生态气候区表现出各自的优势：地处亚热带地区的早实核桃‘香玲’脂肪酸含量丰富(67.4%),脂肪酸(ω-6PUFA/ω-3 PUFA =8.06)、氨基酸均衡性好(SRC=68.18),晚实核桃‘清香’坚果重(单果重14.27g)、出仁率高(53.18%)、氨基酸均衡性好(SRC=70.45);地处暖温带地区的早实核桃‘香玲’坚果重(单果重12.83g)、出仁率高(66.53%)、氨基酸总量(21.84 g/100g)、必需氨基酸含量(5.91 g/100g)丰富,晚实核桃‘清香’则脂肪酸含量丰富(72.41%)且均衡性好(ω-6PUFA/ω-3 PUFA =5.77)。说明不同品种类型的核桃坚果品质受生态气候区气候、果园土壤等环境因素影响存在差异。     

小麦新鲜根系铁氰化钾还原酶与氢、钾跨膜运输的关系

为研究麦根质膜氧化还原反应与钾离子吸收的关系,比较不同基因型小麦吸钾差异,以水培和短期生物吸收试验方法,检测到小麦根系铁氰化钾还原酶的存在,而且不同品种铁氰化钾还原酶还原活性不同。反应液中由于根系内外质子和钾离子的迁移,加与不加铁氰化钾,不同品种根系都引起了溶液pH的改变,但外部电子受体Fe(CN)63-的存在即铁氰化钾还原酶底物的存在加强了溶液pH改变（更多为降低）的程度。铁氰化钾还原也明显影响了根系钾离子的吸收或外排动态,但不同品种影响不同。小麦根细胞膜氧化还原系统明显影响质子和钾离子的跨膜运输,不同品种影响不同。     

棉花脂肪酸合成酶基因GhKAR和GhENR表达载体构建及其功能初探

酮脂酰ACP还原酶(β-ketoacyl-(ACP)reductase,KAR)和烯脂酰ACP还原酶(β-enoyl-(ACP)reductase,ENR)是脂肪酸合成过程中2个关键的还原酶,主要负责脂肪酸碳链的延伸。为明确棉花同源基因的功能,克隆了陆地棉Gh KAR和Gh ENR基因,并构建表达载体;应用花粉管通道法转化棉花,经过PCR检测和Southern杂交,筛选出阳性单拷贝植株,分析其后代株系中目的基因对油分含量以及抗寒性的影响。通过对转基因T4株系含油量的测定,发现转基因株系种子的含油量比野生型对照提高10.2%~14.1%,与对照差异极显著。通过气相色谱法分析转基因株系种子的脂肪酸成分,发现不饱和脂肪酸含量比野生型对照增加4.04%~6.02%。测定4℃低温胁迫下各个转基因株系不同时间生理指标的变化,结果表明:转基因株系的相对电导率和脯氨酸含量积累明显高于对照,而丙二醛含量低于对照,在不同胁迫时间与对照表现出显著或极显著差异,过表达的转基因株系的抗寒性有所提高。     

不同品种稻花鲤肌肉中脂肪酸组成比较分析

对相同养殖条件下的3个不同品种稻花鲤开展其肌肉中脂肪酸组成及含量的测定分析,以期为稻田养殖品系的优化选择提供科学依据。结果表明：福瑞鲤肌肉粗脂肪含量显著低于瓯江彩鲤和本地鲤(P＜0.05);本地鲤、福瑞鲤和瓯江彩鲤3种稻花鲤肌肉中均检测出16种脂肪酸,其脂肪酸组成种类基本相似;亚油酸、油酸、棕榈酸、硬脂酸、棕榈一烯酸和a-亚麻酸的含量占脂肪酸总量的92%以上;不饱和脂肪酸占比分别达到(73.52±0.13)%、(75.67±0.07)%和(75.69±0.19)%;福瑞鲤肌肉中多不饱和脂肪酸和必需脂肪酸的相对含量均显著高于本地鲤和瓯江彩鲤(P＜0.05),表明福瑞鲤稻花鱼具有比本地鲤和瓯江彩鲤稻花鱼更为优良的脂肪酸营养和肉质风味。     

Estimating the fatty acid composition of the oil in intact-seed rapeseed (Brassica napus L.) by near-infrared reflectance spectroscopy

The objective of this work was to evaluate the potential of near-infrared reflectance spectroscopy (NIRS) as a rapid method to estimate the fatty acid composition of the oil in intact-seed samples of rapeseed. A total of 549 samples (3 g intact seed) from selected mutant and breeding lines were scanned by NIRS, and 220 of them were selected and scanned again by using two different adapters, which reduced the sample size to 300 and 60 mg, respectively. Selected samples were analysed by gas liquid chromatography and calibration equations for individual fatty acids were developed. Calibrations for oleic, linoleic, linolenic, and erucic acid were highly accurate, with values of r² in cross validation from 0.95 to 0.98 (samples of 3 g), from 0.93 to 0.97 (300 mg), and from 0.84 to 0.96 (60 mg). Calibrations for palmitic and stearic acid were less accurate, with values of r² in cross validation always lower than 0.8, probably because of the narrow range available for these fatty acids. The accuracy of the calibration equations for eicosenoic acid was very low (r² = 0.69 in 3 g samples), although improved equations were developed (r² from 0.78 to 0.91) when the relationship between erucic and eicosenoic acid was taken into account. We conclude that NIRS is a powerful technique to estimate the fatty acid composition of the oil in rapeseed, provided that samples covering a wide range of fatty acid levels are available, with the advantage that such estimation is possible with few additional costs when NIRS is used for the determination of other seed quality traits. This revised version was published online in July 2006 with corrections to the Cover Date.     

Physiological Investigation of the Impact of Nitrogen and Sulphur Application on Seed and Oil Yield of Rapeseed (Brassica campestris L.) and Mustard (Brassica juncea L. Czern. and Coss.) Genotypes

Field experiments were conducted to determine the physiological basis of the effects of N and S interactions on seed and oil yield of Brassica species. Five combinations of N and S (in kg ha⁻¹) 0S+100N (T₁), 40S+60N (T₂), 40S+100N (T₃), 60S+100N (T₄) and 60S+150N (T₅), were used for this purpose. Nitrate reductase (NR) activity and ATP-sulphurylase activity in the leaves were measured at various growth stages, as the two enzymes catalyse rate-limiting steps of the assimilatory pathways of nitrate and sulphate, respectively. The activities of these enzymes were strongly correlated with seed and oil yield. The highest nitrate reductase activity, ATP-sulphurylase activity and yield were achieved with the combination T₃ in both species. Any variation from this combination decreased the activity of these enzymes, resulting in a reduction of the seed and oil yield of Brassica species. The higher seed and oil yield achieved in these species at T₃ could be due to optimization of leaf soluble protein and photosynthetic rate, as these parameters are influenced by N and S assimilation.