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High-throughput functional gene screening method and system for quantitative analysis of cell phenotype images

A functional gene and quantitative analysis technology, applied in image analysis, material analysis, sequence analysis, etc., can solve the problems of low throughput and inability to accurately quantify cell phenotypes, achieve good repeatability, fast recognition speed, and reduce workload Effect

Active Publication Date: 2021-02-05
HUAZHONG UNIV OF SCI & TECH
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Problems solved by technology

[0005] The present invention solves the technical problem of low flux in cell screening technology in the prior art and cannot accurately quantify cell phenotype, and provides a high-throughput functional gene screening method, thereby greatly improving screening efficiency and improving results The degree of accuracy, and the functional genes can be sorted and classified according to the time point and degree of their influence on the cell phenotype, so as to build a gene interaction network

Method used

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  • High-throughput functional gene screening method and system for quantitative analysis of cell phenotype images
  • High-throughput functional gene screening method and system for quantitative analysis of cell phenotype images
  • High-throughput functional gene screening method and system for quantitative analysis of cell phenotype images

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Embodiment 1

[0042] The 103 phosphorylation regulator genes in the iEKPD database were screened to see whether they were related to autophagy.

[0043] 1. Generate a training set: import GFP-Atg8 into yeast cells to express and produce green fluorescence, use FM4-64 to mark vacuoles to produce red fluorescence, and the conditions for phenotype generation include nitrogen reduction and rapamycin induction (rapamycin induction) Wait. The cells were non-autophagy phenotype cells at 0 hours, and the cells underwent autophagy after 2 hours under reduced nitrogen conditions. Generate large numbers of images of phenotyped and non-phenotyped cells using fully automated fluorescence microscopy.

[0044] 2. Image processing: enhance the fluorescence signal by enhancing the contrast and deconvolution and denoising. Deconvolution and denoising: use PSF (point spread function) of different sizes to restore blurred images, analyze and reconstruct PSF, so as to improve the restored image.

[0045] 3. ...

Embodiment 2

[0060] figure 1 (a) Cell phenotype at 0h after atg1 knockout under nitrogen reduction condition, no autophagy at this time; figure 1 (b) Cell phenotype at 2 hours after atg1 knockout under nitrogen reduction condition, at which point autophagy did not occur; figure 1 (c) is the cell phenotype at 0h after SNF1 is knocked out under nitrogen reduction condition, and there is no autophagy at this time; figure 1(d) Cell phenotype at 2 h after SNF1 knockout under nitrogen reduction condition, at which time autophagy occurs. like figure 1 As shown in (d), GFP-Atg8 enters FM4-64-labeled vacuoles during autophagy; as figure 1 (a), figure 1 (b) and figure 1 As shown in (c), GFP-Atg8 is mainly localized outside the vacuole when cells are not autophagy. We can teach computers to recognize this phenotypic difference to distinguish autophagic from non-autophagic cells.

[0061] figure 2 It is a flowchart of a high-throughput gene screening method based on quantitative analysis of m...

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Abstract

The invention discloses a high-throughput functional gene screening method and system for quantitative analysis of cell phenotype images, and belongs to the technical field of gene screening. The images of cells with phenotypes to be screened and those without phenotypes to be screened are captured by a fully automatic fluorescence microscope; converted into black and white binary images of phenotypes to be screened and without phenotypes to be screened respectively; and then divided into individual phenotypes to be screened The image of the cell and the image containing a single cell without a phenotype to be screened; the corresponding part of the image containing a single cell with a phenotype to be screened in the image of a cell with a phenotype to be screened is used as a positive training set, which will contain a single cell without a phenotype to be screened The corresponding part of the cell image in the cell image without the phenotype to be screened is used as a negative training set, and the final model that can identify the cell phenotype is obtained; the image of the gene knockout or overexpression cell to be screened is identified by the final model, and the sample to be screened is obtained. Screening for association of genes with phenotypes. The method improves screening efficiency and accuracy.

Description

technical field [0001] The present invention relates to the technical field of gene screening, more specifically, to a high-throughput functional gene screening method and system for quantitative analysis of cell phenotype images. Background technique [0002] Many experimentalists are committed to identifying functional genes related to specific cell behaviors and constructing the link between genes and phenotypes, which will help humans interpret genes, understand diseases, and develop drugs. For example, the abnormality of autophagy is closely related to the occurrence of various diseases including cancer. Japanese scientist Yoshinori Ohsumi discovered the key gene regulating autophagy through yeast gene screening experiments. Because of his contribution to autophagy, he was awarded the Nobel Prize in Medicine in 2016. Yoshinori Ohsumi disclosed a technique using SDS-PAGE to detect GFP-ATG8 cleavage. This technique uses the difference in mobility between GFP-ATG8 and GFP...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G06K9/00G06K9/34G06T7/60G16B30/00G01N21/84
Inventor 薛宇宁万山郭亚萍
Owner HUAZHONG UNIV OF SCI & TECH
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