番茄交替氧化酶基因的克隆和表达(英文)

Cloning and Expression of an Alternative Oxidase Gene from Lycopersicon esculentum

利用简并PCR扩增产物做探针筛选番茄cDNA基因文库获得一个全长交替氧化酶cDNA基因LeAoxlau。经序列分析得出,该基因全长1418bp,编码区序列长1077 bp,编码约40 kD的前体蛋白。该蛋白在转运到线粒体时被加工成32kD的成熟蛋白。Southern印迹杂交分析结果显示该基因以单拷贝形式存在于番茄的基因组中RTPCR显示,该基因在在番茄植株的根、茎、叶和子叶中表达。重组表达实验表明该基因能在大肠杆菌中表达。

A full-length cDNA gene(LeAoxlau) was isolated from a cDNA library made from ripening fruit of tomato 'UC-82B'(Lycopersicon esculentum), after probing with alternative oxidase(AOX) gene fragments, obtained by degenerate primer PCR. Sequence analysis showed that LeAoxlau was 1 418 bp long and contained a 1 077-bp open reading frame encoding a about 40 kD precursor protein which is processed to a mature protein of 32 kD. Southern blot analysis suggested LeAoxlau is present as a single copy in the genome of tomato. RT-PCR analysis indicated LeAoxlau was expressed in roots, stems, leaves and cotyledons of tomato plants. A recombinant construct containing the open reading frame sequence of the LeAoxlau was transformed into Escherichia coli to express the alternative oxidase precursor protein.