玉米PEPC基因导入籼稻恢复系及高光效材料的创制
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摘要
水稻(Oryza sativa L)作为重要的粮食作物，其现有高产品种的光能利用率在1％-1.5％，而光能利用率的潜力可达3-5％。本研究以转玉米PEPC基因的粳稻Kitaake(潮霉素作筛选标记)为基因供体，与籼型恢复系蜀恢881、蜀恢527、蜀恢955进行杂交，以期导入PEPC基因改良籼型恢复系光合生理特性，并对玉米PEPC基因导入蜀恢881后的光合作用特性、产量性状进行了系统分析，主要结果如下：
1．用40mg／L的潮霉素培养基对蜀恢881的BC_1F_1代种子进行潮霉素抗性检验，初步筛选出含有玉米PEPC基因的植株。
2．用潮霉素引物对三个不同世代的植株进行扩增的结果表明，玉米PEPC基因已经整合进蜀恢881中，并能够稳定的遗传。
3．通过杂交转育，杂交后代的PEPCase活性得到了极大的提高。转PEPC的蜀恢881、蜀恢527、蜀恢955比原品种高8～20倍，表明在杂交后代中玉米PEPC基因不仅能够稳定的遗传而且得到充分表达。
4．在自然条件下，导入玉米PEPC基因的不同世代植株的光合速率均得到大幅度提高。BC_2F_1代植株的净光合速率达到了12～17.3μmol·m~(-2)·s~(-1)，比蜀恢881的7μmol·m~(-2)·s~(-1)提高了50％；在不同光强及不同CO_2浓度下，净光合速率都比蜀恢881高，表现出利用光能和CO_2的能力得到了提高；光量子效率、CO_2羧化效率分析表明分别比蜀恢881提高了75％和40％，而CO_2补偿点却降低了30μmol·m~(-2)·s~(-1)。表明杂交导入玉米PEPC基因后，植株的光合特性得到了改善。
5．导入PEPC基因后，植株的耐光抑制的能力得到改善。植株剑叶叶片经光抑制处理7天后，仍然保持绿色，而蜀恢881的叶片从叶尖开始变黄，叶绿素含量大幅下降。
6．导入玉米PEPC基因后引起植株性状发生较大的变化。BC_2F_1代植株光合作用功能叶叶面积降低了18％，分蘖和有效分蘖分别比蜀恢881增加了21％，50％；而千粒重增加了17％，单株产量增加了30％。回交后代千粒重增加是充实度提高引起的结果(粒长和粒宽无显著变化，粒厚增加)。这些变化可能是玉米PEPC基因导入蜀恢881引起光合效率提高的结果。
论文还讨论了从遗传背景筛选、酶活与光合性状的相关性以及多种遗传背景下深入研究玉米PEPC基因对提高水稻光合效率的作用及其在水稻超高产育种中的应用。
Rice(Oryza saliva L)is one of the important food crops. But the analysis of photosynthesis efficiency to high yield rice varieties showed that its photosynthesis efficiency was 1-1.5%, while its potential photosynthesis efficiency was 3-5%. In this research, restorer lines Shuhui881, Shuhui527 and Shuhui955, as gene acceptors were crossed with the transgenic rice Kitaake containing the PEPC gene from maize. In this study, photosynthetic characteristics and agronomic traits of Shuhui 881 were systematically analyzed, the main results as below:
? The seeds of BC1F1 were screened on the medium with 40mg/L Hygrimycin for the first selection of the plants obtaining foreign PEPC gene.
? The PCR analysis of F1, BC1F1, BC2F1 hybrids and parents indicated that the PEPC gene of maize had been integrated and could be inherited in the progenies.
? In the plants obtaining maize PEPC gene, the activity of PEPCase was increased evidently compared to the check plants, it was increased as much as 8-20 folds. It could be seen that PEPC gene from maize was not only inherited, but also expressed in the progenies plants.
? The light-saturated photosynthetic rates of BC2F1 plants were nearly 100% higher than that of Shuhui881 under natural conditions; The physiological indexes of the BC2F1 plants were markedly superior to that of Shuui881, increased 75% in quantum efficiency and 40% in the carboxylation efficiency, decreased 30 umo昺~-2晄~-1 in CO2 compensation. It demonstrated that the photosynthetic characteristics had been improved after the introducing of maize PEPC gene.
? In the test of the tolerance to photoinhibition, plants containing the PEPC gene from maize exhibited a slighter photoinhibition than the check plants; After test, the leaves of color of the check plants turned yellow, while the leaves of plants containing the PEPC
gene from maize still kept green; The chlorophyll content in the check plants was greatly decreased.
The analysis of hybrids and Shuhui881 showed that the introducing of PEPC gene had some effects on the agronomic traits .The areas of BC2F1 plants were decreased remarkably, while the 1000-grain weight increased 17%, at the same time, the tiller and the panicle forming rate increased. The analysis of grain shape identified that grain length and grain width were not changed, but grain thickness was increased, so the increase of 1000-grain weight was the results of the improvements of photosynthetic characteristics. So, the improvements of the 1000-grain weight, the tiller, the panicle forming rate and the yield per hill maybe were the result of the introducing of PEPC gene from maize.
In order to identify whether and how did the PEPC gene play a great role on the increase of photosynthetic efficiency, it was very necessary to study the backgrounds of hybrids, the correlation between the activity of PEPCase and photosynthetic characteristics, effects on the other rice materials introducing PEPC gene, such as restorer lines, sterile lines, and the photosynthetic characteristics under different conditions.
1．用40mg／L的潮霉素培养基对蜀恢881的BC_1F_1代种子进行潮霉素抗性检验，初步筛选出含有玉米PEPC基因的植株。
2．用潮霉素引物对三个不同世代的植株进行扩增的结果表明，玉米PEPC基因已经整合进蜀恢881中，并能够稳定的遗传。
3．通过杂交转育，杂交后代的PEPCase活性得到了极大的提高。转PEPC的蜀恢881、蜀恢527、蜀恢955比原品种高8～20倍，表明在杂交后代中玉米PEPC基因不仅能够稳定的遗传而且得到充分表达。
4．在自然条件下，导入玉米PEPC基因的不同世代植株的光合速率均得到大幅度提高。BC_2F_1代植株的净光合速率达到了12～17.3μmol·m~(-2)·s~(-1)，比蜀恢881的7μmol·m~(-2)·s~(-1)提高了50％；在不同光强及不同CO_2浓度下，净光合速率都比蜀恢881高，表现出利用光能和CO_2的能力得到了提高；光量子效率、CO_2羧化效率分析表明分别比蜀恢881提高了75％和40％，而CO_2补偿点却降低了30μmol·m~(-2)·s~(-1)。表明杂交导入玉米PEPC基因后，植株的光合特性得到了改善。
5．导入PEPC基因后，植株的耐光抑制的能力得到改善。植株剑叶叶片经光抑制处理7天后，仍然保持绿色，而蜀恢881的叶片从叶尖开始变黄，叶绿素含量大幅下降。
6．导入玉米PEPC基因后引起植株性状发生较大的变化。BC_2F_1代植株光合作用功能叶叶面积降低了18％，分蘖和有效分蘖分别比蜀恢881增加了21％，50％；而千粒重增加了17％，单株产量增加了30％。回交后代千粒重增加是充实度提高引起的结果(粒长和粒宽无显著变化，粒厚增加)。这些变化可能是玉米PEPC基因导入蜀恢881引起光合效率提高的结果。
论文还讨论了从遗传背景筛选、酶活与光合性状的相关性以及多种遗传背景下深入研究玉米PEPC基因对提高水稻光合效率的作用及其在水稻超高产育种中的应用。
Rice(Oryza saliva L)is one of the important food crops. But the analysis of photosynthesis efficiency to high yield rice varieties showed that its photosynthesis efficiency was 1-1.5%, while its potential photosynthesis efficiency was 3-5%. In this research, restorer lines Shuhui881, Shuhui527 and Shuhui955, as gene acceptors were crossed with the transgenic rice Kitaake containing the PEPC gene from maize. In this study, photosynthetic characteristics and agronomic traits of Shuhui 881 were systematically analyzed, the main results as below:
? The seeds of BC1F1 were screened on the medium with 40mg/L Hygrimycin for the first selection of the plants obtaining foreign PEPC gene.
? The PCR analysis of F1, BC1F1, BC2F1 hybrids and parents indicated that the PEPC gene of maize had been integrated and could be inherited in the progenies.
? In the plants obtaining maize PEPC gene, the activity of PEPCase was increased evidently compared to the check plants, it was increased as much as 8-20 folds. It could be seen that PEPC gene from maize was not only inherited, but also expressed in the progenies plants.
? The light-saturated photosynthetic rates of BC2F1 plants were nearly 100% higher than that of Shuhui881 under natural conditions; The physiological indexes of the BC2F1 plants were markedly superior to that of Shuui881, increased 75% in quantum efficiency and 40% in the carboxylation efficiency, decreased 30 umo昺~-2晄~-1 in CO2 compensation. It demonstrated that the photosynthetic characteristics had been improved after the introducing of maize PEPC gene.
? In the test of the tolerance to photoinhibition, plants containing the PEPC gene from maize exhibited a slighter photoinhibition than the check plants; After test, the leaves of color of the check plants turned yellow, while the leaves of plants containing the PEPC
gene from maize still kept green; The chlorophyll content in the check plants was greatly decreased.
The analysis of hybrids and Shuhui881 showed that the introducing of PEPC gene had some effects on the agronomic traits .The areas of BC2F1 plants were decreased remarkably, while the 1000-grain weight increased 17%, at the same time, the tiller and the panicle forming rate increased. The analysis of grain shape identified that grain length and grain width were not changed, but grain thickness was increased, so the increase of 1000-grain weight was the results of the improvements of photosynthetic characteristics. So, the improvements of the 1000-grain weight, the tiller, the panicle forming rate and the yield per hill maybe were the result of the introducing of PEPC gene from maize.
In order to identify whether and how did the PEPC gene play a great role on the increase of photosynthetic efficiency, it was very necessary to study the backgrounds of hybrids, the correlation between the activity of PEPCase and photosynthetic characteristics, effects on the other rice materials introducing PEPC gene, such as restorer lines, sterile lines, and the photosynthetic characteristics under different conditions.
引文
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