水稻苍白叶突变体pgl3（t）的遗传分析和基因定位
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摘要
叶绿素是地球上最重要的光合色素,植物叶绿素的含量与其光合速率、营养状况及生物产量等密切相关。水稻是单子叶植物发育分子生物学研究的理想模式植物,开展水稻叶绿素相关突变体的基因克隆和功能基因组学研究,不仅为揭示水稻叶绿素合成代谢机制、基因表达调控和叶绿体的发育提供理论依据,对水稻超高产育种也具有重要的应用价值。
本研究组从经EMS处理的粳稻品种日本晴后代中得到一株苍白叶(pale-green-leaf)叶色突变体,该叶色突变体的表型特征是:从苗期开始,整个生育期表现叶色苍白,抽穗后发现穗部也较野生型的略显苍白,突变体的株高和分蘖与野生型植株没有明显区别,结实率有轻微的下降。
取该突变体与野生型日本晴分蘖盛期的功能叶片,采用丙酮法进行叶绿素含量的测定,结果显示,与野生型日本晴相比,突变体pgl3(t)的叶绿素a、b的含量明显下降。对该突变体进行遗传分析表明,该叶色突变是受单隐性核基因控制的。将该突变体与已报道的和本实验已有的其它叶色突变体进行等位性检测,结果各杂交后代都能在表型上互补,表明该基因不与目前已报道的叶色相关基因等位,是一个新的叶色相关的基因,鉴于目前已有pgl2(t)的报道,故将该突变体暂时命名为pgl3(t) (pale-green-leaf3 temporary)。
利用籼稻品种TN1与突变体pgl3(t)杂交得到F1代,F1代自交获得F2分离群体,从F2分离群体中选择突变体表型的单株作为定位群体,共鉴定出1427株苍白叶表型突变单株;根据已公布的分子标记和本实验室检测到的多态性标记,用图位克隆法首先将Pgl3(t)基因初步定位在10号染色体的长臂上。进一步发展新的STS标记、SSR标记和CAPs标记,最终将Pgl3(t)基因精细定位于10号染色体长臂上的两个CAPs标记C2和C3之间,物理距离为64kb,序列分析与基因预测发现该区域含有3个开放与阅读框,目前已正在进行基因序列的预测。
Chlorophyll is one of the most important pigments for photosynthesis. The content of chlo-rophyll is intimately related to the photosynthesis efficiency and nutrition and bio-outputs of the plants. As one of the most important food supplies and the ideal model plants in the study of de-veloped and molecular biology of the monocotyledon, it not only offer theories and evidence for showing us the mechanism of chlorophyll metabolism, regulation of gene expression and the development of chloroplast, it’s also of grate importance in breeding of supper rice.
A leaf color mutant was identified from the descendants of an Indica variety Nipponbare which has been dealt with EMS in our laboratory. The mutant displays a pale green leaf during the whole life stage, with low grain set rate. After several generation growed in Hangzhou and Hainan, we found that the characters of the mutant can be geneticed stably. Genetic analysis showed that the mutant is controlled by a single recessive nuclear gene which is not allelic with the known genes. So it’s a new gene related to leaf colour. The mutation was temporarily desig-nated as pgl3(t)(pale-green-leaf temporary) since there has reported the gene named pgl2. Chlo-rophyll anlysis stated that the contents of both chlorophyll a and b showed a decrease compared with the wild type Nipponbare in the tillering stage.
We used a F2 population generated from the cross between the mutant pgl3(t) and an Indica variety TN1 to map the Pgl3(t) gene. The Pgl3(t) gene was finally located to a 64 kb region be-tween two new designed markers c2 and c3 on the long arm of chromosome 10 after primary, fine mapping. Sequence annotation revealed that there are three ORFs in this 64kb region, all of which had a full length cDNA support. Although we had not confirmed which candidate gene is Pgl3(t), the study would be useful for cloning the Pgl3(t) gene and its function analysis.
本研究组从经EMS处理的粳稻品种日本晴后代中得到一株苍白叶(pale-green-leaf)叶色突变体,该叶色突变体的表型特征是:从苗期开始,整个生育期表现叶色苍白,抽穗后发现穗部也较野生型的略显苍白,突变体的株高和分蘖与野生型植株没有明显区别,结实率有轻微的下降。
取该突变体与野生型日本晴分蘖盛期的功能叶片,采用丙酮法进行叶绿素含量的测定,结果显示,与野生型日本晴相比,突变体pgl3(t)的叶绿素a、b的含量明显下降。对该突变体进行遗传分析表明,该叶色突变是受单隐性核基因控制的。将该突变体与已报道的和本实验已有的其它叶色突变体进行等位性检测,结果各杂交后代都能在表型上互补,表明该基因不与目前已报道的叶色相关基因等位,是一个新的叶色相关的基因,鉴于目前已有pgl2(t)的报道,故将该突变体暂时命名为pgl3(t) (pale-green-leaf3 temporary)。
利用籼稻品种TN1与突变体pgl3(t)杂交得到F1代,F1代自交获得F2分离群体,从F2分离群体中选择突变体表型的单株作为定位群体,共鉴定出1427株苍白叶表型突变单株;根据已公布的分子标记和本实验室检测到的多态性标记,用图位克隆法首先将Pgl3(t)基因初步定位在10号染色体的长臂上。进一步发展新的STS标记、SSR标记和CAPs标记,最终将Pgl3(t)基因精细定位于10号染色体长臂上的两个CAPs标记C2和C3之间,物理距离为64kb,序列分析与基因预测发现该区域含有3个开放与阅读框,目前已正在进行基因序列的预测。
Chlorophyll is one of the most important pigments for photosynthesis. The content of chlo-rophyll is intimately related to the photosynthesis efficiency and nutrition and bio-outputs of the plants. As one of the most important food supplies and the ideal model plants in the study of de-veloped and molecular biology of the monocotyledon, it not only offer theories and evidence for showing us the mechanism of chlorophyll metabolism, regulation of gene expression and the development of chloroplast, it’s also of grate importance in breeding of supper rice.
A leaf color mutant was identified from the descendants of an Indica variety Nipponbare which has been dealt with EMS in our laboratory. The mutant displays a pale green leaf during the whole life stage, with low grain set rate. After several generation growed in Hangzhou and Hainan, we found that the characters of the mutant can be geneticed stably. Genetic analysis showed that the mutant is controlled by a single recessive nuclear gene which is not allelic with the known genes. So it’s a new gene related to leaf colour. The mutation was temporarily desig-nated as pgl3(t)(pale-green-leaf temporary) since there has reported the gene named pgl2. Chlo-rophyll anlysis stated that the contents of both chlorophyll a and b showed a decrease compared with the wild type Nipponbare in the tillering stage.
We used a F2 population generated from the cross between the mutant pgl3(t) and an Indica variety TN1 to map the Pgl3(t) gene. The Pgl3(t) gene was finally located to a 64 kb region be-tween two new designed markers c2 and c3 on the long arm of chromosome 10 after primary, fine mapping. Sequence annotation revealed that there are three ORFs in this 64kb region, all of which had a full length cDNA support. Although we had not confirmed which candidate gene is Pgl3(t), the study would be useful for cloning the Pgl3(t) gene and its function analysis.
引文
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