Map-based cloning of the ALK gene, which controls the gelatinization temperature of rice

As a primary crop in China, rice provides the staple food for more than 50% population in the world. With the increasing grain yield and improvement of living conditions, grain quality has drawn more and more attention. Gelatinization temperature (GT), an important parameter for rice cooking quality, is the critical temperature at which the starch granules start to lose crystallinity and birefringence by irreversible expanding and to change the starch surface from polarized to soluble state c…     

水稻稻米糊化温度控制基因ALK的图位克隆及其序列分析

糊化温度是仅次于直链淀粉含量的评价水稻稻米蒸煮品质的重要指标. 许多文献报道, 糊化温度受一主效基因控制. 采用图位克隆法分离克隆了水稻糊化温度基因(ALK), 序列分析表明其编码可溶性淀粉合酶Ⅱ. 进一步比较不同品种间该基因DNA序列与碱消法分析结果, 推测出ALK基因编码区内的碱基替换可能引起了支链淀粉晶体层结构的改变, 从而导致糊化温度的变化.     

Map-based cloning of the <Emphasis Type="Italic">ALK</Emphasis> gene,which controls the gelatinization temperature of rice

Gelatinization temperature (GT) is an important parameter for evaluating the cooking and eating quality of rice besides amylose content (AC). The inheritance of the genes affecting GT has been widely studied and is considered to be controlled by a major gene. Here, we report the map-based cloning of rice ALK that encodes the soluble starch synthase II (SSSII). Comparison between the DNA sequences from different rice varieties, together with the results obtained with digestion of the rice seeds in alkali solution, indicates that the base substitutions in coding sequence of ALK may cause the alteration in GT.     

无载体主干序列的bar和cecropin B 基因表达框共转化水稻

基因枪等直接转化法可将去除质粒载体主干序列的外源基因表达框导入植物基因组,消除质粒主干序列对植物基因组的影响,同时由于转基因分子较小,比较容易实现多个基因的共转化,在植物基因工程育种上有重要的应用价值,研究了基因枪介导外源基因表达框（包括启动子、基因开放阅读框和终止子）转化水稻的影响因素和转基因的整合模式,结果表明：（1）基因枪介导外源基因表达框转化水稻的频率约在0.1%～0.5%之间,非选择标准基因与选择标记基因的共转化频率约为50%～60%,增加基因表达框DNA浓度可提高转化率。相同基因构建物对不同品质水稻的转化率不同,基因构建物的侧边序列对基因枪转化的品种差异可能具有重要影响。（2）非选择标记基因cecropin B表达框在水稻基因组内整合模式简单,仅有1～3个拷贝;筛选标记基因bar表达框却比完整质粒转化后的整合模式复杂得多,插入拷贝数在4～14个之间,线形基因表达框的游离DNA末端和CaMV 35S启动中子重组热点介导的转基因重组可能是导致bar基因表达框复杂整合的重要原因。     

基因枪转化获得的转基因水稻中外源基因整合与表达规律研究

通过基因枪法将cecropin B和bar基因共转化水稻,获得多个水稻优良品系的转化材料,对转基因的结构和表达的系统分析,发现外源基因的整合模式多种多样,有简单也有复杂,其中插入位点数的变化范围为1－7个,拷贝数的变化范围为1－10个,转基因拷贝数的多少与其表达和沉默不存在必然的联系,相同整合模式的转基因事件中基因的表达存在较大的差异,选择标记基因bar比非选择标记基因cecropin B的表达框的完整性和转录概率要高,但是发现大部分表达框完整的bar基因发生基因沉默,而在终止子发生序列丢失的cecropin B基因的表达则明显提高。