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为了探究铁皮石斛叶色突变机理,以铁皮石斛叶色突变体(doyo1)和正常植株(TP35)为试验材料,对其表型、叶绿素含量进行测定,并进行无参转录组测序分析。结果表明,doyo1与TP35相比,株高、茎粗均较低,生长速度较慢;叶绿素含量下降,仅为TP35的15%左右,叶绿素a/b比值(Chl a/b)显著升高。利用转录组测序技术分别从doyo1和TP35叶片中获得4.58Gb和4.90Gb有效数据(clean data),de novo组装后得到60 363条通用基因(Unigenes),其中长度大于1kb的有12 390条,N50为1 295bp。对Unigenes进行表达分析,共获得1 560条差异表达基因(DEGs)。对得到的DEGs进行生物学功能注释,获得总注释信息为1 325条。通过对注释信息进行GO和KEGG富集分析,结合定量PCR验证,结果表明,铁皮石斛叶色突变体形成的原因可能是do GLK1低水平表达和do Ftsz不表达导致叶绿体合成和分裂受阻,叶片细胞内叶绿体数量大大降低,进而叶绿素(Chl)和类胡萝卜素(Car)含量整体下降。本试验结果为研究铁皮石斛叶色突变体形成的机理提供了依据,同时,为观叶型铁皮石斛定向育种提供了参考。
In order to explore the mechanism of leaf color mutation in Dendrobium candidum, the phenotypes and chlorophyll contents of Dendrobium candidum leaf color mutant (doyo1) and normal plant (TP35) were tested, and the non-reference transcriptome sequencing analysis was carried out. The results showed that, compared with TP35, doyo1 had lower plant height and stem diameter and slower growth rate; chlorophyll content decreased only about 15% of TP35 and Chl a / b ratio increased significantly. Using the transcriptome sequencing technique, 4.58Gb and 4.90Gb clean data were obtained from doyo1 and TP35 leaves, respectively. After de novo assembly, 60363 common genes (Unigenes) were obtained, of which 12 390 were longer than 1kb and N50 For 1 295bp. Unigenes expression analysis, a total of 1 560 differentially expressed genes (DEGs). Biological functions of DEGs were annotated and the total annotation information was 1 325. The GO and KEGG enrichment analysis of annotation information and the validation of quantitative PCR showed that the reason for the formation of leaf color mutant of Dendrobium candidum may be that the low level of do GLK1 and the absence of do Ftsz lead to the obstruction of chloroplast synthesis and division, The number of chloroplasts decreased greatly, and the content of chlorophyll (Chl) and carotenoids (Car) decreased. The results of this study provide a basis for the study of the formation mechanism of Dendrobium candidum leaf color mutant, at the same time, provide a reference for the directional breeding of Dendrobium candidum.