当前位置:技术服务>
PAT-seq

PAT-seq: A method to study the integration of 3-UTR dynamics with gene expression in the eukaryotic transcriptome


A major objective of systems biology is to quantitatively integrate multiple parameters from genome-wide measurements. To integrate gene expression with dynamics in poly(A) tail length and adenylation site, we developed a targeted next-generation sequencing approach, Poly(A)-Test RNA-sequencing. PAT-seq returns (i) digital gene expression, (ii) polyadenylation site/s, and (iii) the polyadenylation-state within and between eukaryotic transcriptomes. PAT-seq differs from previous 3' focused RNA-seq methods in that it depends strictly on 3' adenylation within total RNA samples and that the full-native poly(A) tail is included in the sequencing libraries. Here, total RNA samples from budding yeast cells were analyzed to identify the intersect between adenylation state and gene expression in response to loss of the major cytoplasmic deadenylase Ccr4. Furthermore, concordant changes to gene expression and adenylation-state were demonstrated in the classic Crabtree-Warburg metabolic shift. Because all polyadenylated RNA is interrogated by the approach, alternative adenylation sites, noncoding RNA and RNA-decay intermediates were also identified. Most important, the PAT-seq approach uses standard sequencing procedures, supports significant multiplexing, and thus replication and rigorous statistical analyses can for the first time be brought to the measure of 3'-UTR dynamics genome wide.

2015 Harrison et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

 

(PDF) PAT-seq: A method to study the integration of 3-UTR dynamics with gene expression in the eukaryotic transcriptome. Available from: https://www.researchgate.net/publication/278850445_PAT-seq_A_method_to_study_the_integration_of_3%27-UTR_dynamics_with_gene_expression_in_the_eukaryotic_transcriptome [accessed Dec 28 2018].


可选择性多聚腺苷酸化(alternative polyadenylation,APA)是一种普遍存在于真核生物中的基因调控机制。APA事件的发生能使一个基因产生多种mRNA转录本,从而增加转录本的复杂度。选择不同的多聚腺苷酸化位点可以使不同的转录本具有不同的编码序列,或具有不同长度的3'端非翻译区(3'untranslated region,3'UTR)。不同长度的3'UTR可引起RNA结合蛋白或微小RNA(miRNA)的结合位点发生变化,进而影响mRNA的稳定性、定位和翻译效率。近年来,APA作为调控基因表达的重要方式越来越受到重视。


APA现象广泛存在于真核生物中,绝大多数真核基因具有多个poly(A)位点。水稻是最重要的一种粮食作物,也是一种单子叶模式植物。虽然已有研究在水稻中分析了一些APA,但是APA在发育和组织特化中的作用,仍然是不明确的。在本研究中,研究人员使用Poly(A) Tag测序(PAT-Seq)法,系统地调查了14个不同水稻组织和发育阶段的全基因组APA景观图。

结果表明,APA显著介入了发育和数量性状位点(QTL)基因表达。在所有表达的基因中,有大约48%的基因用APA来产生转录组和蛋白质组多样性。一些基因开关APA位点可让差异表达的基因利用交替3UTR区。有趣的是,成熟花粉中的APA是截然不同的,在那里,一组poly(A)因子存在差异表达水平,APA位点有着不同的分布,从而表明在配子体发育过程中有一个独特的mRNA 3’端形成规律。


同样有意思的是,统计分析表明,QTL往往将APA用于许多农艺性状的基因表达调控,从而指出了APA在水稻生产中的一个潜在重要作用。这些结果为作物中APA的高级分析,提供了迄今为止最全面和高分辨率的资源,并揭示了APA与真核生物性状的形成是如何联系在一起的。

蓝景科信技术有限公司
地址:北京市海淀区高里掌路1号院翠湖科技园4号楼3层西侧

电话:400-618-7099
   

关注微信公众号了解更多信息




备案号:京ICP备14015656号-1 蓝景科信(北京)技术有限公司

留言

QQ

微信

电话

我要留言

邮箱注册 手机号注册
   * 名称:
   * 邮箱:
* 手机号:
* 验证码:
   * 密码:
* 确认密码:
   * 选择:
* 学校名称:
* 专业名称:
* 公司名称:
邮箱验证 手机号验证
   * 邮箱:
* 手机号:
* 验证码:
   * 密码:
* 确认密码: