摘要
【目的】研究mi RNA在香蕉不同器官中的表达,初步探索mi RNA在香蕉生长发育过程中的可能作用。【方法】利用茎环引物的半定量和实时荧光定量PCR策略,检测香蕉中的保守mi RNA,并进一步研究mi RNA在香蕉根系、叶片、雄花和果实组织中的表达差异。【结果】mi RNA156d和mi RNA162a具有相似的表达模式,其最高表达量均出现在雄花组织中。mi R156d在雄花中的表达量(2.25)明显高于根系(0.17)、果实(0.34)和叶片(1.00);mi R162a在雄花中的表达量最高,为4.36,根系组织次之,为1.72,而在叶(1.00)和果实(0.99)中的表达几乎相同。mi R164e在根系中的表达量最高,为2.37,雄花次之,为1.93,而在叶(1.00)和果实(0.98)中表达几乎相同。mi R169h在4种组织中的表达量差异不明显。【结论】以香蕉各组织总RNA为模板,通过以SYBR Green为染料的Stem-loop q RT-PCR方法,成功地在香蕉根系、叶片、雄花和果实4种器官中均检测到了保守mi RNA的表达,表明本研究所建立的体系是定性或定量检测香蕉植株中保守mi RNA的准确而有效的方法。同时,在香蕉的不同组织中发现了具有显著表达差异的mi RNA,符合mi RNA在不同组织的表达活性具有多样性的规律。
[Objective] MicroRNAs （miRNAs） are endogenous non-coding small RNAs （sRNAs） with a wide range of regulatory functions in plant development and stress responses. The banana （Musa acuminata, AAA group） is one of the most important fruit crops in tropical and subtropical countries with a high nutritional value for human health. It comprises an important part in the diet of millions of people around the world. Thousands of miRNAs have been identified in many plant species, whereas only a limited number of miRNAs have been predicted in Musa acuminata （A genome） and Musa balbisiana （B genome）. To obtain more insight into the role of miRNAs in banana growth and development, we implemented conserved miR- NAs express patterns in four tissues. [Methods] Roots, leaves, flowers, and fruits were collected from banana plants （Musa acuminata, AAA group, ＇Brazilian＇） grown at the fruit science experimental station of the Chinese Academy of Tropical Agricultural Sciences （Haikou, Hainan province, China）. After collection, all the samples were immediately frozen in liquid nitrogen and stored at -80 ℃ until use. Total RNA was isolated from different tissues following their reference （Chaiet al., 2014） and briefly exposed to RNAasefree DNAase I. cDNAs were synthesized by M-MLV according to the manufacturer＇s instruction, using the specific stem-loop primers for miRNAs. Stem-loop RT-PCR and qRT-PCR were applied to detect the tissuespecific expression levels of conserved miRNA in banana roots, leaves, flowers, and fruits. After reverse transcription, the products of each reaction were diluted 10 times to avoid potential primer interference in the qRT-PCR reactions. Each reaction consisted of 2 μL of product from the diluted reverse transcription reaction, 2 μL of primers （forward and reverse, 10 μmol. L^-1）, 10 μL of FastStart Universal SYBR Green Master （ROX）, 0.5 μL Reference Dye Ⅱ, and 5.5 μL of nuclease-free water. The reaction was initiated with pre-denaturation at 95 ℃ fo
出处
《果树学报》
CAS
CSCD
北大核心
2016年第7期777-782,共6页
Journal of Fruit Science
基金
国家自然科学基金(31501043
31401843)
"十二五农村领域国家科技计划"(2011AA10020605)
海南省自然科学基金(20153116)
国家现代农业产业技术体系项目"国家香蕉产业技术体系"(CARS-32)
