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Uses and relevant drugs of human RPL34 (ribosomal protein L34) gene

A gene and drug technology, applied in the use of human RPL34 and related drugs, can solve the problems of less understanding of the function of human homologous proteins and less mammals.

Active Publication Date: 2015-02-25
AFFILIATED HOSPITAL CHINA ACADEMY OF MILITARY MEDICAL SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, the current research on this protein is more common in plants and insects [Lan Q, Niu LL, Fallon AM. Mosquito ribosomal protein rpL31 resembles rat rpL34: cDNA and deduced amino acid sequence. Biochim Biophys Acta 1994, 1218: 460-462. Dai Z, Gao J, An K, Lee JM, Edwards GE, An G. Promoter elements controlling developmental and environmental regulation of a tobacco ribosomal protein gene L34. Plant Mol Biol 1996, 32:1055-1065. Devitt ML, Stafstrom JP. Cell cycle regulation during growth-dormancy cycles in pea axillary buds.Plant Mol Biol1995,29:255-265.], less involved in mammals, especially the ability of human homologous proteins to understand less

Method used

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  • Uses and relevant drugs of human RPL34 (ribosomal protein L34) gene
  • Uses and relevant drugs of human RPL34 (ribosomal protein L34) gene
  • Uses and relevant drugs of human RPL34 (ribosomal protein L34) gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0083] Example 1 Preparation of RNAi lentivirus against human RPL34 gene

[0084] 1. Screening for effective siRNA targets against the human RPL34 gene

[0085] Retrieve RPL34 (NM_000995) gene information from Genbank; design effective siRNA targets for RPL34 gene. Table 1 lists one of the effective siRNA target sequences for the RPL34 gene.

[0086] Table 1 siRNA target sequence targeting human RPL34 gene

[0087] SEQ ID NO

TargetSeq

1

TGCTGTAAGACCTAAAGTT

[0088] 2. Preparation of lentiviral vector

[0089] Aim at the siRNA target (taking SEQ ID NO: 1 as an example) to synthesize a double-stranded DNA Oligo sequence (Table 2) with Age I and EcoR I restriction endonucleases at both ends; use Age I and EcoR I restriction enzyme Act on the pGCSIL-GFP carrier (provided by Shanghai Jikai Gene Chemical Technology Co., Ltd., figure 1 ), to linearize it, and identify the digested fragments by agarose gel electrophoresis.

[0090] Table 2 Double-s...

Embodiment 2

[0108] Example 2 Real-time fluorescent quantitative RT-PCR method to detect the silencing efficiency of RPL34 gene

[0109] Human lung cancer H1299 cells in the logarithmic growth phase were digested with trypsin to make a cell suspension (the number of cells was about 5×10 4 / ml) were inoculated in a 6-well plate and cultured until the cell confluency reached about 30%. According to the multiplicity of infection (MOI, H1299:5,) value, an appropriate amount of the virus prepared in Example 1 was added, the culture medium was replaced after 24 hours of cultivation, and the cells were collected after the infection time reached 4 days. Total RNA was extracted according to the instruction manual of Invitrogen's Trizol. According to the M-MLV instruction manual of Promega Company, RNA was reverse-transcribed to obtain cDNA (see Table 7 for the reverse transcription reaction system, react at 42° C. for 1 h, and then bathe in a water bath at 70° C. for 10 min to inactivate reverse t...

Embodiment 3

[0116] Example 3 Detection of proliferation ability of tumor cells infected with RPL34-siRNA lentivirus

[0117] Human H1299 cells in the logarithmic growth phase were trypsinized to make a cell suspension (the number of cells was about 5×10 4 / ml) were inoculated in a 6-well plate and cultured until the cell confluency reached about 30%. According to the multiplicity of infection (MOI, H1299:5), an appropriate amount of virus was added, and the culture medium was replaced after 24 hours of culture. After the infection time reached 4 days, the cells of each experimental group in the logarithmic growth phase were collected. The complete medium was resuspended into a cell suspension (2×10 4 / ml), inoculate a 96-well plate at a cell density of about 2000 / well. 5 replicate wells in each group, 100 μl per well. After laying the board, place at 37°C, 5% CO 2 Incubator cultivation. From the second day after plating, the Cellomics ArrayScan VTI high-content screening analyzer (Th...

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Abstract

The invention discloses uses and relevant drugs of human RPL34 (ribosomal protein L34) gene. The invention discloses the uses of the human RPL34 gene in tumor treatment, tumor diagnosis and drug preparation. The invention further constructs small molecule interfering RNA of the human RPL34 gene, an interfering nucleic acid construction body of the human RPL34 gene, interfering lentivirus of the human RPL34 gene, and discloses the uses thereof. The siRNA (small interfering RNA) or the nucleic acid construction body containing the siRNA sequence and the lentivirus provided by the invention can specifically inhibit the expression of the human RPL34 gene, and particularly the lentivirus can efficiently infect target cells and efficiently inhibit the expression of the RPL34 gene in target cells so as to inhibit the growth of tumor cells and promote the apoptosis of the tumor cells, and has vital significance in the tumor treatment.

Description

technical field [0001] The present invention relates to the field of biotechnology, and more specifically relates to the use of human RPL34 and related medicines. Background technique [0002] RNA interference (RNA interference, RNAi) is an immune response to viral infection. It was first discovered in plants. It mainly refers to the fact that some short fragments of double-stranded RNA can decompose specific mRNA, thereby blocking the expression of this gene. . RNAi is currently an effective tool widely used in functional genomics research, and RNAi can also be used for genetic screening to study the molecular mechanism of cancer. At the same time, it is also used to extensively study the influence of polymorphism of multiple genes on tumor phenotype. Since 1998, a certain gene-specific double-stranded RNA was first introduced into nematode cells, and it was found that it can regulate gene expression [Fire A, Xu S, Montgomery MK, Kostas SA, Driver SE, Mello CC. Potent and...

Claims

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Application Information

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IPC IPC(8): A61K48/00A61K45/00A61P35/00C12N15/113C12N15/867C12N7/01C12Q1/68
Inventor 高红军杨绍兴汤传昊秦海峰刘晓晴王红李晓燕李俭杰王伟霞曹跃琼
Owner AFFILIATED HOSPITAL CHINA ACADEMY OF MILITARY MEDICAL SCI
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