Novel human protein with function for promoting mouse NIH/313 cell transformation and coding sequence thereof

A cell transformation, human protein technology, applied in the direction of anti-animal/human immunoglobulin, organic chemistry, animal/human peptide, etc., can solve the problem of lack of functional gene high-throughput and so on

Inactive Publication Date: 2005-12-07
NEWORGEN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002] Human genomics research is currently a hot spot in the world. In addition to large-scale sequencing of human chromosome DNA and expressed sequence sequencing (EST), there is still a lack of high-throughput methods for screening functional genes starting from function.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] Embodiment 1: Acquisition of cDNA gene and its promoting effect on mouse NIH / 3T3 cell clone formation

[0087] FP17548, FP17581 and FP17780 came from human fetal cDNA libraries constructed by conventional methods; LP2254, LP2261, LP2477, LP2537, LP2561, LP2642, LP2698, LP2709, LP3663 and LP3727 were derived from normal liver cDNA libraries constructed by conventional methods. The method is as follows: take fetal tissue (FP clone) or normal liver tissue (LP clone), use Trizol reagent (GIBCO BRL company) to extract total RNA according to the manufacturer's instructions, and use mRNA purification kit (Pharmacia company) to extract mRNA. The cDNA library of the above mRNA was constructed with the pCMV-script TMXR cDNA library construction kit (Stratagene). The reverse transcriptase was changed to MMLV-RT-Superscript II (GIBCO BRL), and the reverse transcription reaction was carried out at 42°C. Transformed XL 10-Gold competent cells, obtained 1 × 10 6 cfu / μg titer cDNA li...

Embodiment 2

[0090] Example 2: Obtaining full-length gene by PCR from placenta or normal liver cDNA:

[0091] Fetal tissue (FP clone) or normal liver tissue (LP clone) was taken, total RNA was extracted with Trizol reagent (GIBCO BRL company) according to the manufacturer's instructions, and mRNA was extracted with mRNA purification kit (Pharmacia company). Use MMLV-RT-Superscript II (GIBCO BRL) and reverse transcriptase to perform reverse transcription at 42°C to obtain placental or fetal cDNA. Use the specific primers for each gene (as shown in the table below), at 97°C for 3'1 cycles. 94°C 30″60°C 30″72°C 1’35 cycles, 72°C 10’1 cycles for PCR amplification to obtain the amplified products of each protein gene containing the complete open reading frame sequence. The amplified product was verified by sequencing and was consistent with the sequence measured in Example 1, and then the amplified product was transferred to a host cell using conventional techniques to obtain a recombinant pro...

Embodiment 3

[0094] Embodiment 3: cDNA clone sequence analysis

[0095] 1. FP17548

[0096] A: Nucleotide sequence (SEQ ID NO: 1) Length: 2748 bases

[0097] B: Amino acid sequence (SEQ ID NO: 3) Length: 216 amino acids

[0098] C. Nucleotide and amino acid combination sequence (SEQ ID NO: 2) clone number and protein name: FP17548

[0099] Start codon: 1856 ATG Stop codon: 2504 TGA Protein molecular weight: 23817.56

[0100] 2. FP17581

[0101] A: Nucleotide sequence (SEQ ID NO: 4) length: 2070 bases

[0102] B: Amino acid sequence (SEQ ID NO: 6) Length: 113 amino acids

[0103] C. Nucleotide and amino acid combination sequence (SEQ ID NO: 5) clone number and protein name: FP17581

[0104] Start codon: 676 ATG Stop codon: 1015 TGA protein molecular weight: 12295.65

[0105] 3. FP17780

[0106] A: Nucleotide sequence (SEQ ID NO: 7) length: 2392 bases

[0107] B: Amino acid sequence (SEQ ID NO: 9) Length: 140 amino acids

[0108] C. Nucleotide and amino acid combination sequence (S...

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Abstract

The present invention discloses a novel human protein with the function of promoting 3T3 cell transformation, polynucleotide for coding said polypeptide and the method for producing said polypeptide by means of recombination technology. Said invention also discloses the agonist for resisting said polypeptide and its therapeutic action. Said invention also discloses the application of the polynucleotide coding said human protein with the function of promoting 3T3 cell transformation.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular, the invention relates to a new polynucleotide encoding a human protein with the function of promoting 3T3 cell transformation, and a polypeptide encoded by the polynucleotide. The present invention also relates to the use and preparation of such polynucleotides and polypeptides. Background technique [0002] Human genomics research is currently a hot spot in the world. In addition to large-scale sequencing of human chromosomal DNA and expressed sequence sequencing (EST), there is still a lack of high-throughput methods for screening functional genes starting from function. [0003] Cancer is one of the major diseases that endanger human health. In order to effectively treat and prevent tumors, people have paid more and more attention to gene therapy of tumors. Therefore, there is an urgent need in this field to develop and study human proteins and their agonists / inhibitors related to...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07H21/00C07K14/435C07K14/47C07K16/18C12N15/11C12N15/12C12N15/63C12N15/64C12P21/02
Inventor 顾健人杨胜利
Owner NEWORGEN
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