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Gene markers associated with rectal adenocarcinoma

A rectal adenocarcinoma and gene technology, applied in gene therapy, drug combination, biological testing, etc., can solve the problems of poor patient compliance, invasiveness, high cost, low sensitivity and specificity

Active Publication Date: 2019-03-01
QINGDAO MEDINTELL BIOMEDICAL CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the early diagnosis of rectal adenocarcinoma mainly depends on colonoscopy, imaging examination and serological marker carcinoembryonic antigen (CEA). Colonoscopy is the most reliable method for the diagnosis of rectal adenocarcinoma, but it is invasive and expensive. Poor patient compliance; CT and ultrasonography are not easy to find small lesions, and the early diagnosis of rectal adenocarcinoma is limited; CEA is a widely used serum marker in clinical practice, with low sensitivity and specificity, and is mainly used for rectal adenocarcinoma Patient Therapy Monitoring

Method used

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  • Gene markers associated with rectal adenocarcinoma
  • Gene markers associated with rectal adenocarcinoma
  • Gene markers associated with rectal adenocarcinoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Example 1 Screening for gene markers associated with rectal adenocarcinoma

[0060] 1. Collection of samples

[0061] 8 cases of rectal adenocarcinoma non-metastatic tissue and rectal adenocarcinoma metastatic tissue samples were collected, and all the above samples were obtained with the consent of the organizational ethics committee.

[0062]2. Preparation of RNA samples (operated using QIAGEN tissue RNA extraction kit)

[0063] 1) Make ice, take the tissue and put it into a mortar, grind it into fine powder, keep the tissue frozen during the grinding process.

[0064] 2) Transfer the tissue into a 1.5ml EP tube, add 1ml Trizol reagent, grind finely with a grinding rod, fully homogenize with a vortex mixer, and ice-bath for 10 minutes.

[0065] 3) Centrifuge at 12000 rpm for 10 min at 4°C.

[0066] 4) Pipette the upper aqueous phase into another new 1.5ml EP tube, add isopropanol equal to the volume of the supernatant, pipette repeatedly, and ice-bath for 10 minute...

Embodiment 2

[0082] Example 2 QPCR sequencing to verify the differential expression of MANBA genes

[0083] 1. According to the detection results of high-throughput sequencing, the MANBA gene was selected for large-sample QPCR verification. According to the sample collection method in Example 1, 70 cases of non-metastatic rectal adenocarcinoma tissue and 70 cases of metastatic rectal adenocarcinoma tissue were selected.

[0084] 2. The RNA extraction steps are the same as in Example 1.

[0085] 3. Reverse transcription:

[0086] 1) Reaction system:

[0087] reagent

volume

MgCl 2

2μl

10×RT Buffer

1μl

RNase-free water

3.75μl

dNTP mix

1μl

RNase inhibitor

0.25μl

AMV reverse transcriptase

0.5μl

Oligo-dT Adapter Primer

0.5μl

Experimental sample

1μl

[0088] 2) Reverse transcription reaction conditions

[0089] According to the reverse transcription reaction conditions in RNA PCR Kit (A...

Embodiment 3

[0108] Example 3 Inhibition of MANBA gene expression

[0109] 1. Cell culture: human rectal adenocarcinoma cell line SW480, in DMEM medium containing 10% calf serum and 1% P / S at 37°C, 5% CO 2 , Cultivated in an incubator with a relative humidity of 90%. The medium was changed once every 2-3 days, and 0.25% trypsin was used for routine digestion and passage.

[0110] 2. siRNA design

[0111] siRNA sequences targeting MANBA:

[0112] siRNA1-MANBA:

[0113] The sense strand is 5'-AAAUCUGUAGUAAGAAUCCUG-3' (SEQ ID NO.7);

[0114] The antisense strand is 5'-GGAUUCUUACUACAGAUUUAA-3' (SEQ ID NO.8),

[0115] siRNA2-MANBA:

[0116] The sense strand is 5'-UGCUAUAGGUCCAGUUAUCCA-3' (SEQ ID NO.9);

[0117] The antisense strand is 5'-GAUAACUGGACCUAUAGCAAA-3' (SEQ ID NO.10),

[0118] siRNA3-MANBA:

[0119] The sense strand is 5'-AAUAGUGACUUCAUUGAACAG-3' (SEQ ID NO.11);

[0120] The antisense strand is 5'-GUUCAAUGAAGUCACUAUUGG-3' (SEQ ID NO.12)

[0121] Negative control siRNA seque...

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PUM

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Abstract

The invention discloses a genetic marker-MANBA associated with rectal adenocarcinoma and further discloses applications of the MANBA gene in production of products for prejudging risk of rectal adenocarcinoma metastasis, diagnosing transferring of rectal adenocarcinoma and judging recurrence of rectal adenocarcinoma. Meanwhile, the invention discloses an application of the MANBA gene in production of drugs for inhibiting / preventing rectal adenocarcinoma from being transferred.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to the application of MANBA gene in the diagnosis and treatment of rectal adenocarcinoma. Background technique [0002] Rectal adenocarcinoma is one of the common malignant tumors of the digestive tract. In my country, the mortality rate of rectal adenocarcinoma ranks fifth among malignant tumors. Its morbidity and mortality are increasing year by year, seriously endangering human health. Despite the continuous development of diagnosis and treatment technology, the 5-year survival rate of patients after surgery has not improved significantly. The lack of effective diagnostic methods to achieve early diagnosis and treatment of rectal adenocarcinoma and precancerous lesions is one of the main reasons. Rectal adenocarcinoma is a typical epithelial malignant tumor, and adenocarcinoma is the main histological type of rectal adenocarcinoma. Rectal adenomas also become intraepithelial neoplas...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6886G01N33/68G01N33/574A61K45/00A61K48/00A61K31/713A61P35/00
CPCA61K31/713A61K45/00C12Q1/6886C12Q2600/112C12Q2600/118C12Q2600/158G01N33/57446G01N33/68
Inventor 杨承刚任静
Owner QINGDAO MEDINTELL BIOMEDICAL CO LTD
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