Plasma micro-ribonucleic acid (miRNA) marker related with human Hirschsprung's disease and application of miRNA marker

A Hirschsprung's disease, marker technology, applied in the fields of genetic engineering and clinical medicine, can solve problems such as not getting corresponding attention

Active Publication Date: 2012-02-22
NANJING MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the application of miRNA in plasma in the early diagnosis and monitoring of HSCR has not received corresponding attention. If stable specific plasma miRNA related to the onset of HSCR can be found as a biomarker, and a diagnostic and monitoring kit for the corresponding disease can be developed, not only in This field is in the leading position in the world and can create remarkable economic benefits. It will also be a strong impetus to the prevention and treatment of birth defects in my country.

Method used

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  • Plasma micro-ribonucleic acid (miRNA) marker related with human Hirschsprung's disease and application of miRNA marker
  • Plasma micro-ribonucleic acid (miRNA) marker related with human Hirschsprung's disease and application of miRNA marker
  • Plasma micro-ribonucleic acid (miRNA) marker related with human Hirschsprung's disease and application of miRNA marker

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0087] Embodiment 1 Research Object Selection and Grouping Basis

[0088] From July 2009 to July 2011, the inventor collected blood and tissue samples from children with Hirschsprung and normal children with Hirschsprung who met the requirements from Children's Hospital Affiliated to Nanjing Medical University and other hospitals (clinical diagnostic criteria see figure 1 , figure 2 ), from which 100 healthy controls (average age: 89.03±9.0 days) and 100 Hirschsprung patients (average age: 97.5±7.07 days) who met the requirements were selected as Real-time PCR Subjects for detection of miRNA expression. Specific sample classification criteria are as follows:

[0089] Group A: Healthy control group (n=100, 20 people for microarray screening, 40 people for first-stage verification, 40 people for independent population verification):

[0090] 1. Age between 0 and 6 months;

[0091] 2. No digestive system disease;

[0092] 3. No other congenital malformations;

[0093] 4. N...

Embodiment 2

[0100] Embodiment 2 research object pathological diagnosis type

[0101] Hirschsprung is diagnosed by radiological examination, anorectal manometry, and rectal wall histology in children with no meconium passage, delayed meconium passage, constipation, or stool passage by enema. The characteristics of radiological examination including barium enema mainly have the following manifestations: 1. There is a "cone"-shaped transitional separation zone between the lesion segment and the dilation segment; 2. Irregular contraction of the lesion segment; 3. Barium retention; 4. Rectal spasm No expansion; 5 intestinal wall stiffness. Anorectal manometry mainly shows the absence of internal anal sphincter relaxation reflex; the rhythmic contraction of anal canal is significantly reduced; the resting pressure of the rectum and internal sphincter is higher than normal. The histological examination of the rectal wall mainly observed whether there were ganglion cells and the degree of cell d...

Embodiment 3

[0102] Example 3 Taqman miRNA array screening

[0103] Preparation of cDNA samples: a) Take 100 μl plasma; b) Add 900 μl Trizol, shake and mix well, centrifuge at 12,000 rpm at 4°C for 15 minutes, discard the lower layer of waste liquid; c) Add 1.5 times the volume of supernatant absolute ethanol, shake and mix well, Transfer to a spin column, centrifuge at 12,000 rpm for 15 seconds, and discard the lower layer of waste liquid; d) Add 700 μl of RWT buffer to the spin column, centrifuge at 10,000 rpm for 15 seconds, and discard the lower layer of waste liquid. e) Add 500 μl of RPE buffer to the spin column, centrifuge at 10,000 rpm for 15 seconds, and discard the lower layer. f) Repeat e. g) Add the spin column to a new 2ml tube and centrifuge at 10000rpm for 1 minute to remove the RPE buffer. h) Add 50 μl of DEPC-treated water to the column and centrifuge at 12000 rpm to collect RNA. i) Then cDNA is obtained by RNA reverse transcription reaction. The reverse transcription ...

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Abstract

The invention belongs to the fields of gene engineering and clinical medicine, and discloses a plasma micro-ribonucleic acid (miRNA) marker related with a human Hirschsprung's disease and application of the miRNA marker. The marker is selected from multiple kinds of hsa-miR-34b, hsa-miR-31*, hsa-miR-141 and hsa-miR-194. The marker has specificity and sensitivity on the Hirschsprung's disease, canbe used for preparing a reagent for diagnosing or monitoring the Hirschsprung's disease, can avoid invasive diagnosis, can be used for screening and diagnosis at the early stage, can be repeatedly detected, and is easy in dynamic monitoring.

Description

field of invention [0001] The invention belongs to the fields of genetic engineering and clinical medicine, and relates to plasma microRNA (miRNA) markers related to the occurrence of human Hirschsprung's disease and applications thereof. Background technique [0002] Hirschsprung's disease (HSCR), also known as congenital enteric ganglion deficiency, is a common developmental abnormality of the digestive tract characterized by the complete absence of ganglion cells at the end of the intestinal tract. During the development of the nervous system, enteric nerves develop abnormally, which leads to the absence of ganglion cells in the distal intestinal mucosa and submucosa, and the function of the distal intestinal tract is limited. Finally, delayed or no meconium discharge, abdominal distension and intestinal obstruction are the main symptoms. The main clinical symptoms. HSCR ranks second in the occurrence of congenital digestive tract malformations, second only to congenital...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/11C12Q1/68
Inventor 夏彦恺唐维兵王心如陆春城吴炜周志刚李波
Owner NANJING MEDICAL UNIV
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