Detecting of chloroplast circular RNAs in Arabidopsis thaliana
Detecting of chloroplast circular RNAs in Arabidopsis thaliana
复制标题
拟南芥叶绿体环状RNA的检测
DOI：
10.1080/15592324.2019.1621088
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发表时间：
2019-01-01
影响因子：
2.9
通讯作者：
Wan, Yinglang
中科院分区：
文献类型：
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作者：
Liu, Shuai;Wang, Qiaojun;Wan, Yinglang
环状RNA （circRNAs）是真核生物中鉴定出的共价封闭单链RNA分子。本文报道了10个叶绿体基因组（cpDNA）编码环状rna （cp-circRNA）序列，这些环状rna来自3天大的拟南芥幼苗。它们的亲本序列被称为外显子、内含子和基因间基因序列。由于cp-circRNA的表达水平非常低，我们尝试使用数字聚合酶链反应技术来验证它们的存在。其中四个circrna已被确认：ath_circ_476、ath_circ_477、ath_circ_478和ath_circ_480。利用数字PCR技术定量分析了它们在黄化和去黄化幼苗中的表达水平。其中，外显子circRNA ath_circ_476在黄化苗中高表达。其亲本基因细胞色素C组装蛋白（ycf5）的表达趋势与ath_circ_476相似。鉴于ycf5是一个单外显子基因，我们提出ath_circ_476的环状化是通过一个未知的过程发生的，而不是通过基于rna剪接的机制。因此，我们提供了一种可靠的方法来检测低拷贝的cp-circRNAs，并暗示了RNA在叶绿体中的未知循环机制。
ABSTRACT Circular RNAs (circRNAs) are covalently closed single-strand RNA molecules identified in eukaryotes. Here we report 10 sequences of chloroplast genome (cpDNA) encoded circRNAs (cp-circRNA) from 3-day-old Arabidopsis thaliana seedlings via next-generation sequencing. Their parental sequences were referred to as exonic, intronic, and intergenic gene sequences. Due to the very low level of cp-circRNA expression, we attempted to use a digital polymerase chain reaction technique to validate their existence. Four of these circRNAs were confirmed: ath_circ_476, ath_circ_477, ath_circ_478, and ath_circ_480. Their expression levels in etiolated and de-etiolated seedlings were quantified using digital PCR. Of the four, the exonic circRNA ath_circ_476 was highly expressed in etiolated seedlings. Its parental gene, cytochrome C assembly protein (ycf5), has expression trends similar to ath_circ_476. Given that ycf5 is a single-exon gene, we propose that the circularization of ath_circ_476 occurs via an unknown process rather than via RNA-splicing-based mechanisms. Therefore, we provided a reliable method for detecting cp-circRNAs in low copies and implied an unknown circularisation mechanism of RNA in chloroplasts.