Tumor suppressor, coded protein and application thereof

A tumor suppressor and protein technology, applied in the field of application of the gene and encoded protein, can solve the problem of unconfirmed autophosphorylation combination, etc.

Inactive Publication Date: 2005-01-19
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In previous studies, it was found that the NTKL protein has the typical structure of protein kinases, and it was found...

Method used

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  • Tumor suppressor, coded protein and application thereof
  • Tumor suppressor, coded protein and application thereof
  • Tumor suppressor, coded protein and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0093] Acquisition of the mNTKL-BP1 gene

[0094] 1. Construction of "bait" plasmid pDB-Leu-mNTKL

[0095] The cDNA sequence encoding the mouse mNTKL protein was obtained as a positive clone when using the N-terminus of the human ALC1 gene as a bait protein to screen mouse embryos and fetal brain cDNA libraries, and was fused in the pPC86 vector (ProQuest of U.S. Invitrogen Corporation) TM Yeast two-hybrid system) on the AD 3' end, digested and ligated by restriction enzyme SalI / NotI, two fragments of 0.9kb and 2.2kb, firstly connect the 2.2kb fragment with the double-digested recovery fragment of the vector, and then pass SalI Single enzyme digestion, dephosphorylation treatment, insertion of a 0.9kb fragment, identification of the insertion direction, and completion of subcloning the entire coding sequence of mNTKL into the SalI-NotI cloning site of the vector pDB-Leu (Invitrogen Company), thereby constructing a "bait" Plasmid pDB-Leu-mNTKL.

[0096] 2. Autonomous activati...

Embodiment 2

[0102] Acquisition of human hNTKL-BP1 gene

[0103] The following primers were synthesized,

[0104] Forward primer number: HBF,

[0105] Sequence: 5'-CGG ATG AGC TGG GCA GCA GTG TTG G-3' (SEQ ID NO: 5)

[0106] Reverse primer number: HBR,

[0107] Sequence: 5'-ACC AGA ACT TCA TGT GGC CAA AGC A-3' (SEQ ID NO: 6)

[0108] Using Polymerase Chain Reaction (Polymerase Chain Reaction, PCR) technology, using the Marathon-ready cDNA library of human fetal brain tissue (purchased from U.S. Clontech Company) as template amplification, PCR reaction was performed with Advantage TM 2 Polymerase enzyme, reaction conditions according to the American Clontech company Marathon TM cDNA Amplification kit manual operation.

[0109] The amplified product was sequenced to obtain a DNA sequence (SEQ ID NO: 3) containing a complete open reading frame (open reading frame, ORF) with unknown function. According to the inventor's research on the function of the protein encoded by this gene, it w...

Embodiment 3

[0111] Sequence analysis of hNTKL-BP1 gene

[0112] Using the primers designed by us, the human hNTKL-BP1 gene DNA sequence cloned by PCR reaction is 1214bp in length (SEQ ID NO: 3). The protein coding frame sequence is 21-1202, encoding a protein with 394 codons (Fig. 3 and SEQ ID NO: 4), including two coiled coil domains. Compared with the homologous DNA sequence XM_044455 registered in GenBank, the full length of XM_044455 is 2576bp, encoding a deduced open reading frame (from 21bp-1205bp) containing 394 amino acids, and its corresponding protein sequence accession number is: XP_044455.

[0113] The DNA sequence homology of the human hNTKL-BP1 gene and the mouse mNTKL-BP1 gene reaches 80%, and the amino acid sequence of the protein reaches 89%. However, the N-terminus of human hNTKL-BP1 protein encodes 22 more amino acids than that of mouse (Fig. 2).

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Abstract

The invention discloses a human and mice tumor suppressor and coded protein, wherein the gene relates to the regulation and control for encoded proteins and cell cycles, thus is closely related to the occurrence of tumors, the gene, the encoded protein and their derivatives can be used for diagnosis and treatment for tumor diseases caused by cell cycle control imbalance and medicament screening.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular, the invention relates to a new tumor suppressor gene and its coded protein, as well as the application of the gene and the coded protein. Background technique [0002] Previous work found that NTKL protein (N-terminal kinase like protein, N-terminal kinase-like protein) and its homologous proteins are expressed in Drosophila, nematode, Arabidopsis, mouse and human tissues. [0003] In 2000, Liu et al. cloned the cDNA sequence of mouse mNTKL (GenBank accession No.AF276514), and observed that the 105kd protein encoded by it was widely expressed in different tissues of mice, and separated 3T3 by ultracentrifugation -Different parts of L1 adipocytes, the expression of this protein is found in the cytoplasm and low-density components and other small organelles [SimonC.H.Liu, William S.Lane, Gustav E.LienhardCloning and preliminary characterization of a 105kDa protein with an N-terminalkina...

Claims

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Application Information

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IPC IPC(8): A61K38/17A61P35/00C07K14/435C07K16/18C12N15/11C12N15/12C12N15/63C12P21/00
Inventor 霍克克邸玉君
Owner FUDAN UNIV
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