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Application of dcf1 gene to regulation and control over expression of ATP1B1

A technology of ATP1B1 and gene regulation, applied in the application field of expression, can solve problems such as the decline of learning and memory ability

Inactive Publication Date: 2016-09-28
SHANGHAI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Wen et al. found that dcf1 plays an important role in the differentiation of neural stem cells by studying the changes in gene expression before and after the differentiation of neural stem cells in SD rats; at the same time, experiments proved that overexpressing dcf1 in the mouse neural stem cell line C17.2 can maintain the future development of neural stem cells. These results indicate that the dcf1 gene plays an important role in the development of the nervous system. At the same time, previous laboratory studies have shown that the learning and memory abilities of dcf1 knockout mice are reduced

Method used

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  • Application of dcf1 gene to regulation and control over expression of ATP1B1
  • Application of dcf1 gene to regulation and control over expression of ATP1B1
  • Application of dcf1 gene to regulation and control over expression of ATP1B1

Examples

Experimental program
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Embodiment 1

[0016] Example 1: Cellular immunofluorescence

[0017] Put the newborn rat into a beaker containing absolute ethanol. The amount of absolute ethanol should be able to completely submerge the newborn rat. Observe that the newborn rat is completely immobile and start to take the rat brain. ; Carefully take out the neonatal rat brain, place it in a glass dish pre-cooled with water and pre-cooled PBS, and carefully peel off the hippocampus tissue of the neonatal rat with a pointed tweezers under a stereoscope; transfer the neonatal rat hippocampus tissue to a clean 15ml In the centrifuge tube, use a pipette to gently pipette several times; after observing that the tissue is completely broken, centrifuge it at 4℃, discard the supernatant, add an appropriate amount of pre-cooled 1X PBS, pipette and resuspend, repeat the steps three times; The Neuro medium containing 5% FBS growth factor B-27 and double antibody was gently pipetted to resuspend the cells, 20ul medium was added to 20ul t...

Embodiment 2

[0018] Example 2: Western blotting to detect protein expression

[0019] Transfect HEK293 cells according to different groups. After 48 hours, transfer the cells to a clean 1.5ml centrifuge tube. After washing with PBS, add the lysis buffer that has been pre-added with protease inhibitors (add 400ul lysis buffer to a dish with a diameter of 10cm) After 30 minutes, transfer the tissue / cells to a clean 1.5ml ultracentrifuge tube, balance and centrifuge in an ultracentrifuge at 10,000g at 4℃ for ten minutes; 100ul carefully Transfer the supernatant to a clean 1.5ml centrifuge tube, add 25ul 5X SDS Loading Buffer (1ul mercaptoethanol has been added in advance), mix for a while and incubate in a metal bath at 99°C for 10 minutes; sample can be spotted after instant centrifugation. 80V, 30 minutes; 120V, 90~120 minutes (blue Loading dye to the bottom of the glue); cut out the PVDF glue of appropriate size, and after activation with anhydrous methanol, put it in the transfer liquid with...

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Abstract

The invention relates to application of a dcf1 gene to regulation and control over expression of ATP1B1. Two types of mouse detection including a wild type (as reference) and dcf1 knockout are adopted, and it is found that the dcf1 gene has influences on astrocytes through ATP1B1; it is found through cell immunofluorescence observation that after dcf1 knockout is conducted, the sizes of the astrocytes are reduced; then cellular immunofluorescence and co-immunoprecipitation prove the interaction between DCF1 and the ATP1B1 with the function of influencing the sizes of the astrocytes, research on the interaction relationship and the upstream and downstream adjustment relationship between the DCF1 and the ATP1B1is conducted, meanwhile, an RNA interference technology is utilized to lower the expression quantity of the ATP1B1, it is observed that abnormity of the astrocytes is relieved, and the treatment potential of the DCF1 for dysgnosia caused by abnormity of the astrocytes is finally proved.

Description

Technical field [0001] The invention relates to an application of dcf1 gene to regulate the expression of ATP1B1. Background technique [0002] Intellectual disability is a common disease that causes intellectual impairment and weakened adaptive capacity due to developmental disorders of the nervous system. Patients show delayed language learning, memory loss, weak adaptability, etc. About 2-3% of the global population are affected by this disease, and more than a quarter of these patients are due to abnormal gene levels. [0003] Astrocytes are a type of glial cells in the central nervous system. Astrocytes account for about 20%-40% of the total glial cells, which can regulate extracellular ions and chemical environment, support the brain blood barrier, and provide nutrients for nerve tissue. In addition, astrocytes It plays an important role in the development of the nervous system. Its abnormalities will affect the normal development of the nervous system and cause a series of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/569
CPCG01N33/56966G01N33/68G01N2333/46G01N2800/2814
Inventor 文铁桥王董王娇冯瑞丽
Owner SHANGHAI UNIV
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