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Application of estrogen receptor related receptor gamma (ESRRG) variant transcript in auxiliary diagnosis of pituitary adenoma

A pituitary adenoma, transcript technology, applied in the direction of microbial determination/testing, DNA/RNA fragments, biochemical equipment and methods, etc.

Pending Publication Date: 2021-11-09
BEIJING NEUROSURGICAL INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] There is no report on the application of ESRRG abnormal transcripts in the diagnosis of pituitary adenomas, especially prolactin-type pituitary adenomas

Method used

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  • Application of estrogen receptor related receptor gamma (ESRRG) variant transcript in auxiliary diagnosis of pituitary adenoma
  • Application of estrogen receptor related receptor gamma (ESRRG) variant transcript in auxiliary diagnosis of pituitary adenoma
  • Application of estrogen receptor related receptor gamma (ESRRG) variant transcript in auxiliary diagnosis of pituitary adenoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0078] Example 1 Detection of Estrogen Receptor-related Receptor γ (ESRRG) Abnormal Transcripts in Prolactin Pituitary Adenomas

[0079] RT-PCR reaction was performed on 3 samples of normal pituitary gland and 6 samples of prolactinoma pituitary adenoma to detect the existence of abnormal transcripts of ESRRG.

[0080] Total RNA was extracted from the normal pituitary and prolactinoma samples using RNeasy Mini Kit (74106, Qiagen), and reverse-transcribed using High Capacity cDNA Reverse Transcription Kit (4368814, Thermo Fisher, USA). Subsequently, PCR was performed using I-5™ High-Fidelity Master Mix (I5HM, 200MCLAB).

[0081] The thermal cycling protocol is as follows:

[0082] Initial denaturation was performed at 98°C for 2 min, followed by 32 repetitions of a three-step cycle program consisting of 10 s at 98°C (denaturation), 10 s at 59°C (annealing), and 10 s at 72°C (extension), followed by a final at 72°C Extend for 5 minutes.

[0083] The RT-PCR primers for ESRRG a...

Embodiment 2

[0092] Example 2 Detection of Classical and Variant ESRRG mRNA Expression Levels

[0093] PCR primers (Table 1) were designed for variant ESRRG transcripts and classic ESRRG transcripts respectively, and qPCR verification was performed in the above normal pituitary samples and prolactin pituitary adenoma samples, and the RNeasy Mini Kit (74106, Qiagen) was used to extract the above Sample total RNA was reverse transcribed using the High Capacity cDNA Reverse Transcription Kit (4368814, ThermoFisher, USA). Using Power SYBR TM Green PCR Master Mix (4367659, ThermoFisher) was used for qPCR with a total reaction volume of 10 μL. GAPDH was used as an internal control (n=3 per group). GAPDH was used as a reference gene. Detection of mRNA levels was performed on an ABI 7500 system (Applied Biosystems).

[0094] Table 1

[0095]

[0096] The detection results of ESRRG mRNA expression level showed that the expression level of variant ESRRG in variant ESRRG prolactinoma samples ...

Embodiment 3

[0097] Example 3 Construction of Plasmids pDC316-mCMV-ZsGreen-ESRRG16 and pDC316-mCMV-ZsGreen-ESRRG17

[0098] 1. DNA primer design:

[0099] ESRRG16-NheI-F:

[0100] 5'-CTG GCTAGC GCCACCATGTGGCGAGAATGTGATT-3' (SEQ ID NO: 17)

[0101] ESRRG17-NheI-F:

[0102] 5'-CTG GCTAGC GCCACCATGTCAAACAAGATCGAC-3' (SEQ ID NO: 18)

[0103] ESRRG-NotI-R:

[0104] 5'-AGTG GCGGCCGC TCAGACCTTGGCCTCCAACATTTCCAA-3' (SEQ ID NO: 19)

[0105]2. Using plasmid pCDH-ESRRG16-EF1-coGFP-T2A-Puro and plasmid pCDH-ESRRG17-EF1-coGFP-T2A-Puro (purchased from Beijing Li Keli Biotechnology Co., Ltd.) as templates, PCR amplified target genes respectively For ESRRG16 and ESRRG17, NheI and NotI restriction sites were respectively introduced at both ends of the primers. The PCR reaction conditions are shown in Table 1.

[0106] Table 2

[0107]

[0108] Set up the reaction program according to the conditions shown in Table 2 to carry out the PCR reaction.

[0109] table 3

[0110]

[0111] The ...

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Abstract

The invention discloses an application of an estrogen receptor related receptor gamma (ESRRG) variant transcript in auxiliary diagnosis of pituitary adenoma, especially in auxiliary diagnosis of prolactin pituitary adenoma. By detecting whether the ESRRG transcript has an abnormal transcript or not and developing and applying the diagnostic kit, the pituitary adenoma, especially prolactin pituitary adenoma, can be more conveniently and easily diagnosed, so that a patient can know the disease risk in the early stage of the disease, a simple and effective auxiliary means is provided for drug screening, drug effect evaluation and targeted therapy of prolactin pituitary adenoma, especially prolactin pituitary adenoma.

Description

technical field [0001] The invention relates to the field of auxiliary diagnosis of pituitary adenomas, in particular to the application of estrogen receptor-related receptor gamma (ESRRG) mutant transcripts in the auxiliary diagnosis of pituitary adenomas, especially prolactin-type pituitary adenomas. Background technique [0002] Pituitary tumor is one of the most common intracranial tumors, accounting for about 10-15% of intracranial tumors, and the incidence rate of the population is about 8-9 / 10,000. With the development of imaging technology, its detection rate shows a trend of increasing year by year. According to different endocrine characteristics, pituitary tumors can be divided into several subtypes such as prolactin type, growth hormone type, corticotropin type, thyrotropin type, nonfunctional type and mixed type. The tumor secretes various hormones abnormally, leading to significant endocrine disorder symptoms such as infertility, acromegaly, Cushing syndrome, ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12N15/11G01N33/68G01N33/574
CPCC12Q1/6886G01N33/6893G01N33/57484G01N33/57407C12Q2600/156
Inventor 郭靖李储忠谢微嫣王红云龚磊李丹张亚卓
Owner BEIJING NEUROSURGICAL INST
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