A UCR sequence, kit and detection method for detecting high expression in B-cell lymphoma tissue
A lymphoma, B cell technology, applied in the field of tumor molecular biology, to achieve the effects of good stability, good application prospects and high sensitivity
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Embodiment 1B
[0055] Example 1 Analysis of UCR microarray differential expression between B-cell lymphoma tissue and paired normal tissue
[0056] 1. Materials and methods
[0057] 1. Materials
[0058] The tissue samples were obtained from inpatient surgical resection samples of 20 B-cell lymphoma patients (numbered 1-20), 15 of which were paired B-cell lymphoma tissues and adjacent normal tissues.
[0059] 2. Method
[0060] 2.1 Extraction of total RNA from tumor tissue and paired normal tissue
[0061] The total RNA extracted from B-cell lymphomatous tumor tissue and paired normal tissue was operated according to the instructions of Qiagen's RNA extraction kit (RNeasyMicroKit, #74004). The kit contains RNase-FreeDNase I (lyophilized), which can effectively remove the genome Medium DNA. The purity and concentration of the extracted RNA were then quantified using a NanoDrop ND 1000 Nucleic Acid Quantifier (NanoDrop Technologies, Wilmington, Delaware).
[0062] 2.2 Fluorescent labeling...
Embodiment 2
[0081] Example 2 Preliminary verification of the differential expression of uc.189 in B-cell lymphoma tissues and paired paracancerous normal tissues by real-time fluorescent quantitative qRT-PCR
[0082] 1. Experimental materials
[0083] Another 40 pairs (numbered 21-60) of B-cell lymphoma tissues and paired adjacent normal tissues were selected to conduct the first instance verification of the expression difference of uc.189 by qRT-PCR.
[0084] 2. Experimental methods and results
[0085] 1. Primer specificity screening and identification
[0086] According to the uc.189 gene locus, the uc.189-related transcript gene sequence was extracted from the UCSC Genome Brower database, and primers were designed for uc.189 with Primer 5 primer design software;
[0087] (1) The designed primers were evaluated by Oligo7, and 3 pairs of designed primer sequences were obtained;
[0088] The first pair: upstream primer SEQ ID NO.2
[0089] Downstream primer SEQ ID NO.3
[0090] The ...
Embodiment 3
[0124] Example 3 qRT-PCR further verified the differential expression of uc.189 in B-cell lymphoma tumor tissues and paired normal tissues
[0125] 1. qRT-PCR kit composition
[0126] 1.1 Dye-based uc.189PCR kit composition:
[0127] (1) Upstream primer: SEQ ID NO.4
[0128] (2) Downstream primer: SEQ ID NO.5;
[0129] Other reagents refer to SYBR Premix Ex Taq TM II (Tli RNaseH Plus) fluorescence quantitative kit (Code No.RR820A).
[0130] 2. Detection of uc.189
[0131] 2.1 Preparation of total RNA
[0132] Select another 90 pairs of B-cell lymphomatosis (numbered 61-130) tumor tissue and paired normal tissue, according to the ThermoFisher Scientific company's Reagents (Product No. 15596018) Reagents and steps required to extract total RNA, please refer to the instructions for details. The purity and concentration of the extracted RNA were then quantified using a NanoDrop ND 1000 Microvolume UV-Vis Spectrophotometer (NanoDrop Technologies, Wilmington, Delaware).
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