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首页> 美国卫生研究院文献>Plant Physiology >Peach (Prunus persica) endopolygalacturonase cDNA isolation and mRNA analysis in melting and nonmelting peach cultivars.
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Peach (Prunus persica) endopolygalacturonase cDNA isolation and mRNA analysis in melting and nonmelting peach cultivars.

机译：桃（Prunus persica）内聚半乳糖醛酸酶cDNA分离和mRNA分析分别用于融化和未融化的桃子品种。

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摘要

Two distinct partial cDNAs, PRF1 and PRF3, similar in sequence to previously described polygalacturonases, were amplified from ripe peach (Prunus persica L. Batsch cv Flavorcrest) fruit cDNA by the polymerase chain reaction. PRF1-related RNA was present in fruit from early ripening at levels not detected by northern analysis. PRF3-related RNA was readily detectable in ripe fruit by northern analysis. PRF3 was used to isolate a cDNA with a complete open reading frame, PRF5, from a lambda ZAP II cDNA library prepared from poly(A)+ RNA of ripe peach fruit. PRF5 coded for a predicted protein of 393 amino acids with a molecular mass of 41,500 D. The derived amino acid sequence of PRF5 included a putative leader sequence of 23 amino acids, followed by a sequence that matched the N terminus of endopolygalacturonase protein purified from ripe peach fruit. By northern analysis, PRF3-related RNA was undetectable in firm, unripe Flavorcrest fruit. It appeared at low levels as a 1.7-kb transcript in fruit that had begun to ripen and soften and was very abundant in ripe fruit that had undergone the "melting" stage of softening. The marked increase in PRF3-related RNA levels took place over a period of less than 2 d at 20 degrees C and coincided with the climacteric peak in ethylene evolution. Levels of 1-aminocyclopropane-1-carboxylate oxidase-related RNA increased during ripening at a much earlier stage than levels of PRF3-related RNA. Lower levels of 1.7-kb RNA transcript were detected by PRF3 in ripe fruit of the melting cultivar Fragar, which are firmer than Flavorcrest fruit. In ripe fruit of the nonmelting cultivar Carolyn, PRF3 detected a 1.45-kb RNA transcript that was present at low levels. Transcripts of a peach polygalacturonase-related genomic sequence were not detected in ripening fruit.
机译：通过聚合酶链反应从成熟的桃子（Prunus persica L. Batsch cv Flavorcrest）果实cDNA中扩增出两个序列不同的部分cDNA，即PRF1和PRF3，其序列与先前描述的多半乳糖苷酶相似。早熟果实中存在与PRF1相关的RNA，RNA分析未检测到该水平。通过Northern分析很容易在成熟果实中检测到PRF3相关的RNA。 PRF3用于从由成熟桃果实的poly（A）+ RNA制备的λZAP II cDNA文库中分离具有完整开放阅读框PRF5的cDNA。 PRF5编码393个氨基酸的预测蛋白，分子量为41,500D。PRF5的衍生氨基酸序列包括一个推定的23个氨基酸的前导序列，其后的序列与从成熟的纯化的内聚半乳糖醛酸酶蛋白的N末端匹配桃果实。通过Northern分析，在牢固的未成熟的Flavourcrest果实中未检测到PRF3相关的RNA。它在开始成熟和软化的果实中以1.7-kb的转录物低水平出现，并且在经历了“熔化”软化阶段的成熟果实中含量很高。 PRF3相关RNA水平的显着增加发生在20摄氏度下少于2 d的时间内，并且与乙烯进化中的更年期峰相吻合。 1-氨基环丙烷-1-羧酸氧化酶相关的RNA的水平在成熟期间比PRF3相关的RNA的水平要早得多。通过PRF3在融化品种Fragar的成熟果实中检测到较低水平的1.7-kb RNA转录物，该果实比Flavorcrest果实牢固。在非融化品种卡罗琳（Carolyn）的成熟果实中，PRF3检测到1.45kb RNA转录本的含量较低。在成熟的果实中未检测到桃多聚半乳糖醛酸酶相关基因组序列的转录本。

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