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Macrobrachium rosenbergii estrogen-related receptor gene ERR and application thereof

A technology of macrobrachium rosenbergii and receptor gene, which is applied in the field of genetic engineering, can solve the problems of unfavorable wide use, complicated operation of yeast two-hybrid system, etc., and achieves the effects of low cost, convenient source and increased gene expression.

Active Publication Date: 2016-06-01
海南永贺生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the operation of the yeast two-hybrid system is relatively complicated, which is not conducive to widespread use

Method used

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  • Macrobrachium rosenbergii estrogen-related receptor gene ERR and application thereof
  • Macrobrachium rosenbergii estrogen-related receptor gene ERR and application thereof
  • Macrobrachium rosenbergii estrogen-related receptor gene ERR and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] 1. Extraction of total RNA from Macrobrachium rosenbergii

[0035] Take healthy Macrobrachium rosenbergii, anesthetize it in an ice bath for about 2 minutes, dissect and sample, and immediately separate the ovarian tissue. Trizol reagent method was used to extract the total RNA of Macrobrachium rosenbergii, and its OD 260 / 280 =1.91, the electrophoresis result ( figure 1 ) Shows that 28SrRNA and 18SrRNA have clear bands, and the brightness of 28S bands is twice that of 18S, indicating that the total RNA obtained is not contaminated by protein, phenol and genomic DNA and has high purity.

[0036] 2. Synthesis of the first strand of cDNA

[0037] 1) Take out 2μg of the preserved Macrobrachium rosenbergii RNA, add 1μL of polythymidine Oligo (dT) (10mM), mix well and place in a water bath at 70°C for 5 minutes;

[0038] 2) After placing on ice for 5 minutes, add dNTPMixture (10mM) 2μL, 5XRTBuffer 4μL, Rnase inhibitor 1μL (10U / μL), Rnase-freeddH to the EP tube on ice. 2 O8μL, ReverTr...

Embodiment 2

[0059] Tissue expression of ERR gene in Macrobrachium rosenbergii.

[0060] Semi-quantitative PCR was used to detect the expression of the ERR gene of Macrobrachium rosenbergii in various tissues, and the results are as follows Image 6 As shown in the results, the results show that ERRmRNA is highly expressed in the heart, brain, and ovary; the expression in hepatopancreas, testis and gills is weak; no expression signal can be detected in muscle and eyestalk. It shows that ERR is involved in regulating the expression of neuromodulation and other endocrine regulation genes in Macrobrachium rosenbergii, enriching the system signal pathway theory of endocrine regulation in shrimps, and can provide a certain theoretical basis for the research of related shrimp sex regulation.

Embodiment 3

[0062] The expression changes of ERR gene and its protein in Macrobrachium rosenbergii during NP bathing.

[0063] Distribute healthy female Macrobrachium rosenbergii in the early stage of sexual differentiation and maturation into 4 independent plastic buckets (50L) of the same specifications. After one week of domestication, add NP (nonylphenol) to make the final concentration of 0, 5, 25, 125μg respectively / L. Feed commercial feeds on time at 07:00 and 17:00 every day. Keep the water temperature and light conditions the same. After one week of treatment, they are anesthetized with alcohol, sacrificed, collected ovarian samples, treated with liquid nitrogen, and stored in an ultra-low temperature refrigerator at -80℃ save. The collected ovarian samples were subjected to total RNA extraction, reverse transcription, and fluorescence quantitative detection of ERR gene expression changes.

[0064] The result is Figure 7 As shown in Figure 7 The results in showed that: compared w...

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Abstract

The invention discloses a macrobrachium rosenbergii estrogen-related receptor gene ERR. The nucleotide sequence of the gene ERR is shown as SEQ ID NO:1, and the amino acid sequence of the gene ERR is shown as SEQ ID NO:2. The invention further discloses application of the recombined macrobrachium rosenbergii estrogen-related receptor gene ERR to environmental internal-secretion interfering-substance detection and pollution assessment, and macrobrachium rosenbergii ERR protein has important application value in the aspects of natural water environment internal-secretion interfering-substance (like NP) detection and pollution assessment.

Description

Technical field [0001] The invention belongs to the technical field of genetic engineering, and specifically relates to a Macrobrachium rosenbergii estrogen-related receptor gene ERR and its application. Background technique [0002] The estrogen-related receptors ERRα and ERRβ were first discovered by Giguere in 1988. These two new human genes were screened by low-stringency hybridization against the conserved sequence of the DNA binding domain (DBD) of ERα. The estrogen receptor (ER) has a strong homology, hence the name. At present, it has been found that there are three members of the ERR family, namely ERRα (NR3B1), ERRβ (NR3B2) and ERRγ (NR3B3). Together with ER, they constitute a subfamily of the third type of nuclear receptors. Unlike ER, they are not compatible with females. It binds to hormones and does not respond to the stimulation of natural estrogen such as estradiol. Research suggests that ERR can act with coactivators to activate the expression of target genes in...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/12C12N15/70C07K14/72
CPCC07K14/43509C07K14/721
Inventor 朱春华陈华谱赵苗鑫李广丽邓思平洪越群洪宇键
Owner 海南永贺生物科技有限公司
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