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Human protein with function of promoting 3T3 cell conversion and its coding sequence

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

Inactive Publication Date: 2003-03-19
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] PP11646, PP11647, PP12994, PP13187, PP13850 and PP14296 were obtained by constructing a human placental cDNA library by conventional methods; FP852, FP938, FP1047, FP2025, FP2461 and FP3059 were obtained by constructing human fetal cDNA libraries by conventional methods. Take 3, 6, and 9-month-old placental tissue (PP clone) or fetal tissue (FP 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 lib...

Embodiment 2

[0090] Embodiment 2: PCR obtains the expression of full-length gene and recombinant protein from placenta or fetal cDNA

[0091] The placental tissue or fetal tissue of 3, 6, and 9 months old was taken, and the total RNA was extracted with Trizol reagent (GIBCO BRL company) according to the manufacturer's instructions, and the mRNA was extracted with the 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), and perform 3'1 cycles at 97°C. 94°C 30″, 60°C 30″, 72°C 1’35 cycles, 72°C 10′ 1 cycle 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 into a ...

Embodiment 3

[0093] Embodiment 3: cDNA clone sequence analysis

[0094] 1. PP11646

[0095] A: Nucleotide sequence (SEQ ID NO: 1) length: 1527 bases

[0096] 1 GTTTTCCCAG AGTGTTTTTT GGTTTCTGAA AGGCTGAACA CAGGCAGAGT GAGGGAAGGA

[0097]61 TGAAGTCTAC AGGCCATTGT GATGCAGCCC TTCCAGCAGG TGTGACAGAT TATATAAGGA 121 CCATCTGGAA AAGAAACATG CTGCCTGTTG TCTGGAAACC TTCCACTGAC ACCTTCAGAT 181 CAGCAATGTC TTATTTTTCA TTTTACCGCC CATCTGCATG TGCTTGTTTC GTCAGTATGC 241 AACATGCTTC AACAGTGGCA TCTACTTAAT CTGGACTCTT TTGGTTGTAG TGGGAATTGG 301 ATCCGTCTAC TTCCATGCAA CCCTTAGTTT CTTGGGTCAG ATGCTTGATG AACTTGCAGT 361 CCTTTGGGTT CTGATGTGTG CTTTGGCCAT GTGGTTCCCC AGAAGGTATC TACCAAAGAT 421 CTTTCGGAAT GACCGGGGTA GGTTCAAGGT GGTGGTCAGT GTCCTGTCTG CGGTTACGAC 481 GTGCCTGGCA TTTGTCAAGC CTGCCATCAA CAACATCTCT CTGATGACCC TGGGAGTTCC 541 TTGCACTGCA CTGCTCATCG CAGAGCTAAA GAGGCATGAG AGGAACCAGC GAAGGAGACA 601 CAGGAAAGGT GGCCAGCAAG GAGGTGGAGA CAAGGTCTGA CGATGAGTGA CTC...

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Abstract

The present invention discloses one kind of human protein with 3T3 cell conversion promoting function, polynucleotides encoding the polypeptide and the recombinant process of producing the polypeptide. The present invention also discloses the agonist resisting the polypeptide and its treatment effect. The present invention also discloses the application of the polynucleotides encoding the human protein with 3T3 cell conversion promoting function.

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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IPC IPC(8): C07H21/00C07K14/435C07K16/18C12N15/10C12N15/11C12N15/12C12N15/63C12N15/64C12P21/02
Inventor 顾健人杨胜利
Owner NEWORGEN
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