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验证miR-21,miR-31,miR-34A和miR-203的变化通过量化聚合酶链反应在孤立的角质细胞中

Constanza Contreras-Jurado^1,2,3

¹Instituto de Investigaciones Biomédicas Sols-Morreale (IIBM), Consejo Superior de Investigaciones Científicas (CSIC)-Universidad Autónoma de Madrid (UAM), Madrid, Spain. scontjur@uax.es.

Methods in molecular biology (Clifton, N.J.)

|November 23, 2024

概括

这项研究验证了在小鼠角质细胞中通过使用茎环实时PCR进行RNA测序检测到的microRNA (miRNA) 变化. 该方法准确地测量成熟的miRNA水平,帮助研究甲状腺激素信号传递.

科学领域:

分子生物学分子生物学
遗传学是一个遗传学.
内分泌学在内分泌学.

背景情况:

微RNAs (miRNAs) 是细胞功能的关键调节者,调节失调与各种疾病有关.
甲状腺激素 (TH) 对许多生理过程至关重要,其信号传递受到组织中多个miRNA的影响.
需要准确的方法来验证通过RNA测序等高通量技术识别的miRNA表达变化.

研究的目的:

描述和验证一种干环实时PCR方法,用于量化成熟miRNA的表达.
评估野生型和双甲状腺激素受体淘汰 (dTRKO) 鼠的角质细胞中miRNA水平的变化.

主要方法:

利用干环RT原料,在单次反应中高效地合成特定miRNA的cDNA.
测量了从四天大的野生型和dTRKO小鼠中分离的角质细胞中的成熟miRNA表达水平.
使用实时PCR进行miRNA表达量的定量分析.

主要成果:

茎环实时PCR方法成功量化了成熟的miRNA水平.
该研究提供了一种经过验证的方法来确认通过RNA测序观察到的miRNA表达变化.

结论:

茎环实时PCR是验证miRNA表达的可靠方法,特别是研究miRNA参与甲状腺激素信号传递.
这种技术适用于角质细胞,可以用于研究dTRKO小鼠等遗传模型中的miRNA角色.

关键词:

双重甲状腺受体淘汰赛小鼠皮肤表面皮肤的皮肤.角质细胞 (keratinocytes) 是一种细胞.逆向转录是一种反向转录.树干循环实时PCR实时PCR 甲状腺激素受体 (TR) 是一种微RNAs 是一个微型RNA.

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相关概念视频

MicroRNAs

MicroRNAs

MicroRNA (miRNA) are short, regulatory RNA transcribed from introns—non-coding regions of a gene—or intergenic regions—stretches of DNA present between genes. Several processing steps are required to form biologically active, mature miRNA. The initial transcript, called primary miRNA (pri-mRNA), base-pairs with itself forming a stem-loop structure. Within the nucleus, an endonuclease enzyme, called Drosha, shortens the stem-loop structure into hairpin-shaped pre-miRNA. After the pre-miRNA ends...

MicroRNAs

MicroRNAs

MicroRNA (miRNA) are short, regulatory RNA transcribed from introns (non-coding regions of a gene) or intergenic regions (stretches of DNA present between genes). Several processing steps are required to form biologically active, mature miRNA. The initial transcript, called primary miRNA (pri-mRNA), base-pairs with itself, forming a stem-loop structure. Within the nucleus, an endonuclease enzyme, called Drosha, shortens the stem-loop structure into hairpin-shaped pre-miRNA. After the pre-miRNA...