Inhibition of protein degradation increases the Bt protein concentration in Bt cotton

Bacillus thuringiensis(Bt)cotton production is challenged by two main problems,i.e.,the low concentration of Bt protein at the boll setting stage and the lowest insect resistance in bolls among all the cotton plant’s organs.Therefore,increasing the Bt protein concentration at the boll stage,especially in bolls,has become the main goal for increasing insect resistance in cotton.In this study,two protein degradation inhibitors(ethylene diamine tetra acetic acid(EDTA)and leupeptin)were sprayed on the bolls,subtending leaves,and whole cotton plants at the peak flowering stage of two Bt cultivars(medium maturation Sikang 1(SK1))and early maturation Zhongmian 425(ZM425)in 2019 and 2020.The Bt protein content and protein degradation metabolism were assessed.The results showed that the Bt protein concentrations were enhanced by 21.3 to 38.8%and 25.0 to 38.6%in the treated bolls of SK1 and ZM425 respectively,while they were decreased in the subtending leaves of these treated bolls.In the treated leaves,the Bt protein concentrations increased by 7.6 to 23.5%and 11.2 to 14.9%in SK1 and ZM425,respectively.The combined application of EDTA and leupeptin to the whole cotton plant increased the Bt protein concentrations in both bolls and subtending leaves.The Bt protein concentrations in bolls were higher,increasing by 22.5 to 31.0%and 19.6 to 32.5%for SK1 and ZM425,respectively.The organs treated with EDTA or/and leupeptin showed reduced free amino acid contents,protease and peptidase activities and significant enhancements in soluble protein contents.These results indicated that inhibiting protein degradation could improve the protein content,thus increasing the Bt protein concentrations in the bolls or/and leaves of cotton plants.Therefore,the increase in the Bt protein concentration without yield reduction suggested that these two protein degradation inhibitors may be applicable for improving insect resistance in cotton production.

Animal Safety Test of Bacillus thuringiensis BT Protein

[Objectives]To determine the biological safety of BT protein from Bacillus thuringiensis(Bt)fermentation broth to mammals at high doses.[Methods]Healthy mice were randomly divided into 4 groups with 10 mice in each group.The experimental groups were fed with Bt fermentation supernatant at 0.2,0.6 and 1.0 mL/kg,respectively,once a day for 7 consecutive days.The blank control group was fed normally without intragastric administration.[Results]There was no significant difference in blood routine and blood biochemical analysis between the experimental group and the control group.After intragastric administration,the mice were dissected,and no obvious pathological changes were found;the heart,liver,spleen,lung and kidney were taken to make tissue sections,and no pathological changes were found by microscopic observation.[Conclusions]Mice can tolerate high doses of BT protein from B.thuringiensis fermentation broth.

Expression of Bt Protein in Transgenic Pest-resistant Rice

[Objective] The aim of this study was to study on expression of Bt protein in transgenic pest-resistant rice. [Method] Enzyme-linked immunosorbent assay （ELISA） was used to measure Bt protein expression in different tissues of transgenic pest-resistant rice at same growth stage. [Result] Absolute content of Bt protein from high to low was as follows： leaves 〉 immature seeds and glumes 〉 roots 〉 stems in different tissues of transgenic rice in grain-filling stage; Bt protein content of trans- genic rice changed a little in different growth stages （including tillering stage, booting stage, and grain-filling stage）; in general, its level declined a little in later growth stage, but the resistibility would not be influenced significantly. [Conclusion] The ex- periment is significant for pest prevention and transgenic rice breeding.

The effects of high temperature level on square Bt protein concen-tration of Bt cotton

Higher boll worm survival rates were detected after high temperature presented during square period in Bt cotton. The objective of this study was to investigate the effects of high temperature level on the Bt efficacy of two different types of Bt cotton cultivars at squaring stage. During the 2011 to 2013 cotton growth seasons, high temperature treatments ranged from 34 to 44°C in climate chambers, and field experiments under high temperature weather with various temperature levels were conducted to investigate the effects of the high temperature level on square Bt protein concentration and nitrogen metabolism. The climate chamber experiments showed that the square insecticidal protein contents reduced after 24 h elevated temperature treatments for both cultivars, whereas significant declines of the square insecticidal protein contents were detected at temperature 〉38°C, and only slightly numerical reductions were observed when temperature below 38°C. Similar high temperature responses were also observed at the two field experimental sites in 2013. Correspondingly, high temperature below 38°C seems have little effect on the square amino acid concentrations, soluble protein contents, glutamic-pyruvic transaminase（GPT） and glutamic-oxalacetic transaminase（GOT） activities as well as protease and peptidase activities; however, when the temperature was above 38°C, reduced soluble protein contents, enhanced amino acid concentrations, decreased GPT and GOT activities, bolstered protease and peptidase activities in square were detected. In general, the higher the temperature is（〉38°C）, the larger the changes for the above compound contents and key enzymes activities of the square protein cycle. The findings indicated that the unstable insect resistance of the square was related to high temperature level during square stage.

Differential Expression of Bt Protein in Transgenic Bt Cotton

[Objective] The paper was to study the temporal and spatial dynamics of Bt protein expression in transgenic Bt cotton and to determine the inner relationship of Bt protein expression and transgenic Bt cotton. [Method] With transgenic cotton cultivar（ GK19） as the test material,Bt protein contents in different organs,main stem functional leaves at different growth stages and different positions of main stem leaves at different growth stages were studied by enzyme-linked immunosorbent assay. [Result] There were differences in Bt protein content among different organs of transgenic Bt cotton; the Bt protein content of leaves at seedling stage was the highest,followed by flowers,bubs and bolls,and those of roots and stems were relatively low. The Bt protein content of main stem function leaves gradually decreased with the progressing development. There were great differences in Bt protein content among different positions of main stem leaves at different growth stages; the Bt protein content of the 1^（st）-7^（th） top leaves at seedling stage and full budding stage gradually decreased,while those at full flowering stage and full bolling stage first slowly increased then gradually stabilized. [Conclusion] Bt protein expression was found in all organs of transgenic cotton at all growth stages,and the expression level presented temporal and spatial dynamics.

Relationship Between Leaf C/N Ratio and Insecticidal Protein Expression in Bt Cotton as Affected by High Temperature and N Rate

Expression of insecticidal protein for transgenic Bacillus thuringiensis （Bt） cotton is unstable and related to nitrogen metabolism. The objective of this study was to investigate the relationship between leaf carbon nitrogen ratio （C/N） and insecticidal efficacy of two Bt cotton cultivars. C/N ratio and Bt protein content were both measured at peak square period and peak boll period respectively under 5-7 d high temperature and different nitrogen fertilizer rates on the Yangzhou University Farm and the Ludong Cotton Farm, China. All plants were grown in field. The results showed that the C/N ratio enhanced slightly and the Bt protein content remained stable at peak square period, but significant increases for the C/N ratio and decreases markedly for the leaf Bt protein concentration were detected at the peak boll period. The similar patterns at the two growth periods were found for the leaf C/N ratio and Bt protein content by different N fertilizer treatments. When nitrogen rate was from 0 to 600 kg ha-l, the C/N ratio was reduced by 0.017 and 0.006 for Sikang 1 and Sikang 3 at peak square period, compared to the 1.350 to 1.143 reduction for Sikang 1 and Sikang 3 at peak boll period, respectively. Correspondingly, the leaf Bt protein contents were bolstered by 2.6-11.8 and 26.9-36.9% at the two different growth periods, respectively. The results suggested that enhanced C/N ratio by high temperature and nitrogen application may result in the reduction of inseetiocidal efficacy in Bt cotton, especially in peak boll period.

Study on Spatial-temporal Dynamics of Bt Toxic Protein Expression in Insect-resistant Transgenic Cotton and Its Degradation in Soil

[Objective] This study aimed to investigate the spatial-temporal dynamics of Bt toxic protein expression in insect-resistant transgenic cotton and its degradation in soil. [Method] Btcry1Ac toxic protein expression in roots, stems and leaves of transgenic cotton Guoshen GK45 at different developmental stages and the annual average content of BtCry1Ac toxin protein in the topsoil, rhizosphere soil and following cotton-growing area were explored and analyzed by using enzyme linked immuno sorbed assay (ELISA). [Result] The content of exogenous BtCry1Ac toxin protein decreased during the growth process of insect-resistant transgenic cotton; to be specific, the content of BtCry1Ac toxin protein in cotton stems and leaves decreased more slowly and always maintained a high level, while that in roots decreased rapidly and reached a minimum level to the following plant growth and development stage. BtCry1Ac toxin protein was detected in topsoil of both non-transgenic and transgenic cotton-growing areas, and the content of BtCry1Ac toxin protein increased in topsoil of following cotton-growing area, which was very low in rhizosphere soil. [Conclusion] Determination of Btcry1Ac toxic protein provides scientific basis for the risk assessment of the cultivation of genetically modified crops and the safety evaluation of soil ecosystem.

基于转录组分析的低铁胁迫下Bt棉苗杀虫蛋白表达量下降机制

研究低铁对Bt棉杀虫蛋白含量的影响及其生理和分子机制,可为Bt棉抗虫性安全表达提供理论参考。本文以转基因抗虫棉品种中棉50为试验材料,设置正常铁(CK,20.0µmol L^(–1))和低铁(LI,0.1µmol L^(–1))2个铁水平处理,采用水培法研究了低铁胁迫下Bt棉苗期Bt杀虫蛋白含量变化及氮代谢生理特征,通过转录组测序挖掘差异表达基因及其相关代谢途径,并采用qRT-PCR方法对测序结果进行验证。结果表明,与对照处理相比,低铁胁迫下Bt棉根和叶片中杀虫蛋白含量显著下降,且根中杀虫蛋白含量下降幅度更大。低铁胁迫降低了叶片中NH_(4)^(+)-N和NO_(3)^(–)-N含量,降低根和叶片中可溶性蛋白含量和全氮含量。根和叶片中硝酸还原酶、亚硝酸还原酶和谷氨酸合成酶活性的变化趋势与Bt杀虫蛋白含量表现一致。转录组结果表明,根和叶片中分别鉴定出11,661个和8972个差异表达基因,其中有1652个差异表达基因在根和叶片中均下调表达。GO注释表明,低铁处理根和叶片中差异基因的功能都主要富集于刺激反应、细胞壁、质膜、结合、氧化还原活性等。KEGG富集分析显示,苯丙烷类物质生物合成、苯丙氨酸代谢、半胱氨酸和蛋氨酸代谢、激素信号转导、玉米素生物合成、氮代谢、淀粉和蔗糖代谢、丙氨酸、天冬氨酸和谷氨酸代谢和酪氨酸代谢等途径在根和叶片中均发生显著变化。低铁胁迫下与氮素还原、同化途径相关的NR、NiR1和GLT1基因表达显著下调。以上结果表明,低铁胁迫会抑制Bt棉氮代谢相关基因的转录水平,减弱氮代谢的生理活性,抑制Bt杀虫蛋白的合成。

基于Bt毒素的杀虫蛋白理性设计与创新应用策略

Bt毒素是源于苏云金芽孢杆菌(Bacillusthuringiensis)的具有特殊杀虫功能的大分子蛋白,其制剂和转基因作物已广泛用于害虫防治,产生了巨大的经济和社会生态效益。围绕Bt毒素挖掘和提升其应用价值是持续研究的热点,特别是随着Bt毒素结构功能和作用机制日趋明晰,为其功能修饰和创新应用创造了条件,相关研究蓬勃发展,成效显著。大量研究表明,采用定点突变、结构域替换或融合以及抗独特型抗体模拟等策略,是理性设计活性更高、稳定性更强、杀虫谱更广、非靶标生物安全性更高甚至是可用于害虫抗药性治理的有别于母体Bt毒素的突变体、结构杂合体乃至功能效应物抗体等新型杀虫蛋白的有效手段;此外,采用催化毒素活化、驱动毒素靶向受体结合、促进毒素表达以及同源或异源杀虫材料复配或共表达的协同促效等创新增效策略,也是助推Bt毒素应用价值的重要手段。本文总结了Bt毒素结构功能和作用机制,梳理了基于Bt毒素功能修饰的突变体、结构杂合体以及功能效应物抗体等新型杀虫蛋白理性设计和基于Bt毒素功能增效的创新应用策略等相关研究进展,并结合作者团队在模拟Bt毒素杀虫功能效应物抗体靶向设计研发方面的最新成果,探讨了基于Bt毒素的杀虫蛋白理性设计与创新应用策略未来发展动向及潜在可行捷径,为相关研究提供较为全面的最新有价值的文献资料和启发思路。

低温胁迫对Bt棉纤维中杀虫蛋白含量及氮代谢的影响

本研究以常规种泗抗1号(Sikang 1,SK1)和杂交种泗抗3号(Sikang 3,SK3)为材料进行盆栽试验,研究了不同低温水平及其处理持续时间对Bt棉盛铃期纤维中杀虫蛋白含量变化及氮代谢生理特征。结果表明,纤维中的杀虫蛋白含量随着温度的降低总体呈下降趋势,同时低温处理持续期显著影响杀虫蛋白含量。与对照相比,纤维中杀虫蛋白含量的降低幅度随低温胁迫时间的延长而增大。此外,随着处理温度的降低,可溶性蛋白含量、谷丙转氨酶活性、谷草转氨酶活性呈下降趋势,游离氨基酸含量、肽酶活性、蛋白酶活性呈上升趋势,且在低温处理48 h后,均与杀虫蛋白含量呈极显著相关。因此,低温胁迫促使了蛋白质的合成功能下降,分解能力增强,导致可溶性蛋白含量下降,游离氨基酸含量升高,最终导致杀虫蛋白含量下降,且其受低温胁迫持续期显著影响。

Amino acids application enhances flowers insecticidal protein content in Bt cotton

Background:Low insecticidal protein expression at reproductive organs affect insect resistance in Bt transgenic cotton.In order to enhance flower insecticidal protein expression,the conventional cultivar Sikang1(S1)and the hybrid cultivar Sikang3(S3)were used as experimental materials;the applications of selected 5 types of amino acids and 21 types of amino acids were sprayed on the flowers in 2016 and 2017 cotton growing seasons.Results:The flower Bt protein contents increased significantly under the two amino acid treatments in both cultivars,the Bt protein concentration increased by 15.2 to 25.8%compared with the control.However,no significant differences were detected between the two treatments of amino acid application.Increased amino acid and soluble protein contents,enhanced GPT,GOT,protease,and peptidase activities were observed under the amino acid application at the flowering stage.Conclusions:These results suggest that exterior application of the amino acids treatments could bolster the flower insecticidal protein expression.

Impacts of Environmental Factors on Degradation of CrylAb Insecticidal Protein in Leaf-Blade Powders of Transgenic Bt Rice

The determination of the environmental fate of Bt insecticidal protein released by Bt rice plants in paddy soils is a key issue in its ecological risk assessment. In this study, the impacts of soil water content, pH, and temperature on the degradation of CrylAb protein expressed in the leaves of Bt rice KMD2 were studied in the laboratory. Three types of paddy soils were used, i.e., blue clayey paddy soil, pale paddy soil on quaternary red soil, and marine-fluvigenic yellow loamy paddy soil. Ground powders of KMD2 leaf blades were mixed with each type of soil, and degradation dynamics of Cry lAb were measured using enzyme-linked immunosorbent assay （ELISA）. The degradation rate of CrylAb was high at the early experimental stage, but slowed down steadily at middle and later stages, which could be described by exponential equations, with the half-life period of degradation determined as 1.8-4.0 d. The soil water content, pH, and temperature could affect the degradation of CrylAb, but the effects of soil pH and temperature were relatively greater. In general, CrylAb degradations were slower under lower soil pH and temperature conditions, especially for marine-fluvigenic yellow loamy paddy soil.

Dynamics of Bt cotton Cry1Ac protein content under an alternating high temperature regime and effects on nitrogen metabolism

This study was conducted to investigate the effects of alternating high temperature on CrylAc protein content on Bt cotton cultivars Sikang 1 （SK-1, a conventional cultivar） and Sikang 3 （SK-3, a hybrid cultivar）. In 2011 and 2012, cotton plants were subjected to high temperature treatments ranging from 32 to 40℃ in climate chambers to investigate the effects of high temperature on boll shell insecticidal protein expression. The experiments showed that significant decline of the boll shell insecticidal protein was detected at temperatures higher than 38℃ after 24 h. Based on the results, the cotton plants were treated with the threshold temperature of 38℃ from 6：00 a.m. to 6：00 p.m. followed by a normal temperature of 27℃ during the remaining night hours （DH/NN） in 2012 and 2013. These treatments were conducted at peak boll growth stage for both cultivars in study periods of 0, 4, 7, and 10 d. Temperature treatment of 32℃ from 6：00 a.m. to 6：00 p.m. and 27℃ in the remaining hours was set as control. The results showed that, compared with the control, after the DH/NN stress treatment applied for 7 d, the boll shell CrylAc protein content level was significantly decreased by 19.1 and 17.5% for SK-1 and by 15.3 and 13.7% for SK-3 in 2012 and 2013, respectively. Further analysis of nitrogen metabolic physiology under DH/NN showed that the soluble protein content and the glutamic pyruvic transaminase （GPT） activities decreased slightly after 4 d, and then decreased sharply after 7 d. The free amino acid content and the protease content increased sharply after 7 d. The changes in SK-1 were greater than those in SK-3. These results suggest that under DH/NN stress, boll shell CrylAc protein content decline was delayed. Reduced protein synthesis and increased protein degradation in the boll shell decreased protein content, including Bt protein, which may reduce resistance to the cotton bollworm.

Degradation of Cry1Ab Protein Within Transgenic Bt Maize Tissue by Composite Microbial System of MC1

Environmental safety issues involved in transgenic plants have become the concern of researchers, practitioners and policy makers in recent years. Potential differences between Bt maize（ND1324 and ND2353 expressing the insecticidal Cry1Ab protein） and near-isogenic non-Bt varieties（ND1392 and ND223） in their influence on the composite microbial system of MC1 during the fermentation process were studied during 2011-2012. Cry1Ab protein in Bt maize residues didn＇t affect characteristics of lignocellulose degradation by MC1, pH of fermentation broth decreasing at initial stage and increasing at later stage of degradation. The quality of various volatile products in fermentation broth showed that no signifi cant difference of residues fermentation existed between Bt maize and non-Bt maize. During the fermentation MC1 efficiently degraded maize residues by 83%-88%, and cellulose, hemicelluloses and lignin content decreased by 70%-72%, 72%-75% and 30%-37%, respectively. Besides that, no consistent difference was found between Bt and non-Bt maize residues lignocellulose degradation by MC1 during the fermentation process. MC1 degraded 88%-89% Cry1Ab protein in Bt maize residues, and in the fermentation broth of MC1 and bacteria of MC1 Cry1Ab protein was not detected. DGGE profi le analyses revealed that the microbial community drastically changed during 1-3 days and became stable until the 9th day. Though the dominant strains at different fermentation stages had signifi cantly changed, no difference on the dominant strains was observed between Bt and non-Bt maize at different stages. Our study indicated that Cry1Ab protein did not infl uence the growth characteristic of MC1.

渍涝与干旱对不同转Bt基因抗虫棉的影响

对 3个转 Bt基因棉花品种的研究表明 ,花铃期田间渍涝显著降低皮棉产量和棉株内的 Bt蛋白含量。积水后棉花主茎功能叶和幼蕾的 Bt蛋白含量平均分别比对照降低 2 7.7%和 2 9.9% ,主茎下部老叶的 Bt蛋白含量仅降低 6 .6 % ,说明渍涝对生长旺盛器官 Bt蛋白合成的影响远远大于衰老器官。干旱对棉株内 Bt蛋白合成的抑制更强 ,当花铃期干旱使棉叶致萎时 ,主茎下部老叶、幼蕾和主茎功能叶的 Bt蛋白含量分别降低了52 .9%、3 7.0 %和 3 0 .0 %。为充分发挥和利用转Bt基因棉花品种的产量与抗虫潜力 ,棉花生长发育的关键时期应尽量避免缺水干旱 ,及时排除田间积水 。

Bt玉米抗虫蛋白表达的时空动态及其土壤降解研究

采用ELISA定量法对比研究了美国和中国 4种Bt玉米品种 (34B2 4、NK5 8 D1、R× 6 0 1RR/YG和农大 6 1)不同器官Bt蛋白表达的季节动态 ;应用室内 2 5℃恒温模拟分解试验对比了 4个Bt玉米品种秸秆分解释放的Bt蛋白的土壤降解规律。结果表明 ,4种Bt玉米不同器官不同生育期Bt蛋白含量表现出较大的差异 ,总体趋势为随生育期的延长Bt蛋白的表达量降低 ,农大 6 1后期表达量衰退不明显。室内模拟分解试验显示 ,4种Bt玉米秸秆中的Bt蛋白在水中的降解过程呈现初期大量释放 ,前期大量快速降解和后期极低量稳定 3个阶段 ,农大 6 1、34B2 4、NK5 8 D1分解 7d时含量达到最大 ,但R× 6 0 1RR/YG分解 14d达最大。 4种Bt玉米秸秆中的Bt蛋白在室内 2 5℃恒温条件下均能在土壤中快速降解 ,降解过程均呈现前期负指数大量快速降解和中后期极少量稳定两个阶段 ,其DT50 依次为NK5 8 D1(2 .0 9d) <R× 6 0 1RR/YG(2 .6 1d) <农大 6 1(2 .73d) <34B2 4 (3.4 3d) ,DT90 为农大 6 1(10 .5 3d)<R× 6 0 1RR/YG(11.0 9d) <34B2 4 (13.5 2d) <NK5 8 D1(14 .6 7d)。农大 6 1秸秆中ICP在灭菌与未灭菌土壤中降解的对比发现 。

高温胁迫对Bt棉叶片杀虫蛋白表达量和氮代谢影响的研究

通过对两个不同类型Bt棉品种(系)不同生育期高温胁迫及持续时间研究表明,高温胁迫(37℃)48h对盛蕾期和盛花期叶片Bt杀虫蛋白含量影响不大。但在结铃盛期,高温使Bt杀虫蛋白含量明显下降。48h的持续高温结果进一步表明,科棉1号36h前下降快,新洋822于36h后下降快。高温使蛋白酶和肽酶活性提高,丙酮酸转氨酶活性下降,但氨基酸含量增加,说明可溶性蛋白分解增强。

入侵云南草地贪夜蛾种群对5种常用Bt蛋白的敏感性评价

草地贪夜蛾Spodoptera frugiperda(J. E. Smith)是原分布于美洲大陆热带和亚热带地区的一种重要玉米害虫。在当地,种植抗虫转基因玉米是防控草地贪夜蛾危害的主要手段。该虫于2019年1月入侵我国云南省,为明确入侵我国云南的草地贪夜蛾种群对常用Bt蛋白的敏感性水平,本文通过饲料表面涂抹法测定了瑞丽草地贪夜蛾幼虫对Cry1Ab、Cry1Ac、Cry1F、Cry2Ab以及Vip3A等5种Bt蛋白的敏感性。结果表明:几种常用Bt蛋白对瑞丽草地贪夜蛾致死作用顺序为Vip3A>Cry1Ab>Cry1F>Cry2Ab>Cry1Ac,对草地贪夜蛾抑制生长发育的顺序为Cry1Ab>Cry1F>Vip3A>Cry1Ac>Cry2Ab。此外,与美国相对敏感种群比较,云南瑞丽草地贪夜蛾种群对Cry1Ab、Cry1Ac、Cry1F、Cry2Ab和Vip3A的敏感性指标在0.28～3.76之间,表明该入侵种群对此5种Bt蛋白均未产生抗性。此研究可为将来建立以转Bt基因玉米作为防控草地贪夜蛾的技术体系提供依据。

转Bt基因棉杀虫蛋白含量时空分布及对棉铃虫产生抗性的影响

采用ＥＬＩＳＡ法（酶链免疫法）、室内初孵棉铃虫生测法和田间棉铃虫为害调查方法，研究和分析了中国构建的 Ｂｔ（Ｂａｃｉｌｌｕｓ ｔｈｕｒｉｎｇｉｅｎｓｉｓ）基因抗虫棉品系ＧＫ３和美国构建的Ｂｔ基因棉品系新棉３３Ｂ的不同生育期以及花铃期不同器官的杀虫蛋白含量、校正死亡率和田间受害率变化趋势以及它们之间的关系。结果表明，转Ｂｔ基因棉Ｂｔ杀虫蛋白含量在棉花生育过程中呈时空动态变化，在时间分布上，各生育期顶尖平展叶表现为：初花期＞蕾期、花铃期＞苗期＞吐絮期；在花铃期，各器官表现为：功能叶、茎尖＞小蕾、幼铃＞花蕊、花瓣、苞叶、老叶。室内生测幼虫校正死亡率与棉株Ｂｔ杀虫蛋白含量高度一致；田间表现与Ｂｔ杀虫蛋白含量有一定的差异，除主要受Ｂｔ杀虫蛋白影响外，棉铃虫取食选择性，以及残存高龄幼虫为害和棉株各部位营养结构等也影响其抗虫性，造成后期棉铃虫主要为害花、蕾和幼铃，两材料抗虫性表现较为一致。

施氮肥对盐胁迫下Bt棉生长和叶片Bt蛋白含量的影响

以Bt棉品系K638为材料,在盆栽条件下研究了施氮肥对不同程度盐胁迫(土壤含盐量:0(CK)、0.15%(轻度胁迫)和0.3%(中度胁迫))下棉株生长、氮素吸收、Bt蛋白含量和Bt蛋白氮占全氮量比例的影响;同时,在水培条件下研究了不同形态氮素(硝态氮和铵态氮)对NaCl胁迫下Bt蛋白含量的效应。结果表明,氮肥与盐胁迫对棉叶Bt蛋白含量有显著的互作效应。非盐和低盐胁迫下,施氮肥促进了棉株生长(生物量分别提高了3.0和2.8倍)、Bt蛋白合成(分别提高41.0%和90.9%)和全N向Bt蛋白N的转化(分别提高9.3%和15.6%);中度盐胁迫下,施氮肥虽也促进了棉株生长(1.4倍),并提高了叶片全N含量(98.8%)和Bt蛋白含量(83.3%),但并未提高Bt蛋白N占全N量的比例。无论盐胁迫与否,施NO3--N处理的生物量和叶片全氮含量都显著高于施NH4+-N的处理,但由于盐胁迫下NO3--N降低了Bt蛋白N占全氮的比例(11.0%),叶片Bt蛋白含量则略低于NH4+-N处理。据此认为,盐胁迫下施氮肥通过促进棉株对N素的吸收积累并影响全氮转化为Bt蛋白的比例,进而影响Bt蛋白含量。