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Oligonucleotide chip for detecting transplanted kidney function and its uses

A technology of oligonucleotides and oligonucleotide probes, applied in the field of oligonucleotide chips for detecting transplanted kidney function, can solve the problems of high price, heavy workload, and failure to detect, so as to ensure sensitivity and specificity speed, save time and money, and reduce disruptive effects

Inactive Publication Date: 2008-02-13
ZHEJIANG UNIV
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Problems solved by technology

However, it is unrealistic to widely use expression profiling arrays to detect thousands of genes in clinical disease research. Researchers hope that the number of genes studied on the arrays can be reduced, because the data provided by many genes that are not related to the research objects will interfere with the analysis. Interfering with genes related to research subjects, providing useless or erroneous information
And low-abundance and low-expression genes are easily covered by high-abundance expression genes and cannot be detected
At present, there are two main types of gene chips used for clinical research on organ transplantation. One is a high-density short oligonucleotide chip synthesized in situ represented by Affymetrix, which requires high technical requirements and is expensive. Commercially synthesized gene chips, It is not possible to provide customized services according to the needs of researchers; the second is the cDNA chip represented by the research group of Stanford University, which is synthesized by PCR method and then sampled. improvement

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  • Oligonucleotide chip for detecting transplanted kidney function and its uses

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Embodiment 1

[0018] Embodiment 1 Preparation and hybridization of oligonucleotide chip of the present invention

[0019] 1. Chip Preparation

[0020] Referring to Fig. 1, the oligonucleotide chip is composed of a matrix nylon membrane 1, a sampling area 2, and modified and hybridized oligonucleotide probes 3 fixed on the surface of the nylon membrane in a matrix distribution. The oligonucleotide probe 3 has Sequence of SEQ ID NO 1. Specific oligonucleotide probes 3 are respectively spotted on the spotting area 2 of the chip, and the spots are formed into 96 (12×8) matrices, each matrix includes 15 (5×3) spots, and the diameter of the spot is 0.6mm , with a dot pitch of 1.5 μm. According to the existing relevant knowledge, 449 immune-related genes were screened out and synthesized by BIO BASIC INC (Canda) using the 70 bp long oligonucleotide sequence provided by Operon Company, see the sequence of SEQ ID NO 1. The oligonucleotide synthesized at 1 OD was dissolved in 100ul double distille...

Embodiment 2

[0023] Embodiment 2 The application of the oligonucleotide chip prepared by the present invention

[0024] 1: The repeatability and accuracy of the chip

[0025] Pre-experiment 10 probes, the results of two independent hybridizations for each probe, R 2 The value is 0.88-0.95, according to the correlation coefficient R of the chip-to-chip data 2 >0.81 is recognized as the standard of good repeatability of the experiment, and the hybridization data obtained in this paper has good repeatability and can be used for subsequent analysis.

[0026] Compared with the acute rejection group and the stable renal function group, the 10 most significantly differentially expressed genes were screened out. The results of real-time quantitative PCR verification were consistent with the detection results of the microarray. They all showed high expression in the acute rejection group, which verified the microarray hybridization results. accuracy.

[0027] 2: Detection of clinical specimens ...

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Abstract

The invention provides a transplanted renal function test nucleotide chip, consisting of a matrix nylon membrane (1), a spotting area (2) and an oligonucleotide probe (3), which is fixed to the surface of the nylon membrane (1) and is distributed in a matrix way after modification and hybridization. The oligonucleotide probe (3) is equipped with the sequence of SEQ ID NO 1. And a contact-type printing is adopted. Synthesized by BIO BASIC INC (Canda) company, a 70bp oligonucleotide fragment, the specific of which represents one gene, is spotted on the nylon membrane. With a reasonable design, the invention can test the transplanted renal function early, rapidly, cheaply and precisely. And the invention can be applied to the transplanted renal function test.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and specifically relates to an oligonucleotide chip capable of detecting transplanted kidney function, which can detect different transplanted kidney function states including stable state of renal function, acute rejection, acute tubular necrosis and the like. Background technique [0002] Acute rejection is the most common complication after kidney transplantation, which can directly lead to the loss of transplanted kidney function or affect the long-term survival of the transplanted kidney. At present, the diagnosis of acute rejection mainly relies on renal biopsy, which is an invasive examination and is subject to sampling restrictions. Many researchers have tried to find biomarkers of acute rejection and diagnose acute rejection with various experimental methods, but none of them have been widely used clinically. [0003] Chip technology is a cutting-edge molecular biology technology de...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/00G01N33/50C12Q1/68
Inventor 陈江华何强茅幼英杨浩董海涛
Owner ZHEJIANG UNIV
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