Weretilnyk E, Orr W, White T C, Iu B, Singh J
Plant Research Centre, Agriculture Canada, Ottawa, Ontario.
Plant Physiol. 1993 Jan;101(1):171-7. doi: 10.1104/pp.101.1.171.
A cDNA clone, pBN115, encoding a low-temperature-regulated transcript in winter Brassica napus has been isolated. Northern blot analyses show that levels of transcripts hybridizing to pBN115 increase within 24 h of exposure of B. napus to low temperature, peak at 3 d, and then remain at an elevated level for the duration of the cold treatment (up to 10 weeks). Transferring plants from 2 degrees C to room temperature results in the loss of detectable transcripts hybridizing to pBN115 within 1 d. The transcript was not detected in RNA isolated from roots of cold-acclimated B. napus. Results of in vivo labeling of nascent RNA in leaf discs of B. napus with thiouridine suggest that regulation of expression may be transcriptional, at least at the onset of cold temperature. Although drought stress leads to a slight increase in transcript level at room temperature, neither a brief exposure to elevated temperatures nor exogenous application of abscisic acid resulted in the appearance of the transcript represented by pBN115. Furthermore, transcripts hybridizing to pBN115 were present at the same levels whether the plants were acclimated in the light or dark. Hybridization experiments show that pBN115 hybridizes strongly to cold-regulated transcripts in Arabidopsis thaliana, Descurania sophia, and spring B. napus, all of which are cruciferous plants capable of cold acclimation. No hybridizing transcript could be detected in cold-acclimated Spinacea oleracea, winter Secale cereale, or cold-grown Nicotiana tabacum. DNA sequence analysis of pBN115 reveals a single open reading frame that potentially encodes a protein of 14.8 kD. This size closely approximates that of a polypeptide produced by in vitro transcription/translation experiments. Two additional cDNA clones, pBN19 and pBN26, with divergent 5'-and 3'-untranslated regions, were also isolated and found to encode similar, but not identical, polypeptides.
已分离出一个编码冬油菜中低温调节转录本的cDNA克隆pBN115。Northern杂交分析表明，与pBN115杂交的转录本水平在甘蓝型油菜暴露于低温的24小时内增加，在3天时达到峰值，然后在冷处理期间（长达10周）保持在升高水平。将植物从2℃转移到室温会导致在1天内与pBN115杂交的可检测转录本消失。在从冷驯化的甘蓝型油菜根中分离的RNA中未检测到该转录本。用硫尿苷对甘蓝型油菜叶盘新生RNA进行体内标记的结果表明，表达调控可能是转录水平的，至少在低温开始时是这样。尽管干旱胁迫导致室温下转录本水平略有增加，但短暂暴露于高温或外源施用脱落酸均未导致pBN115所代表的转录本出现。此外，无论植物是在光照还是黑暗中驯化，与pBN115杂交的转录本水平相同。杂交实验表明，pBN115与拟南芥、野豌豆和春油菜中的冷调节转录本强烈杂交，所有这些都是能够进行冷驯化的十字花科植物。在冷驯化的菠菜、冬黑麦或冷生长的烟草中未检测到杂交转录本。pBN115的DNA序列分析揭示了一个单一的开放阅读框，其可能编码一个14.8 kD的蛋白质。这个大小与体外转录/翻译实验产生的多肽大小非常接近。还分离出另外两个cDNA克隆pBN19和pBN26，它们具有不同的5'和3'非翻译区，并且发现它们编码相似但不相同的多肽。
