Related Experiment Video
Updated: Aug 9, 2026
MISSION LentiPlex Pooled shRNA Library Screening in Mammalian Cells
Published on: December 21, 2011
Human natural resistance-associated macrophage protein 2: gene cloning and protein identification
1Center for Gene Research, Yamaguchi University, 1144 Kogushi, Yamaguchi, Ube, 755-8505, Japan. fkishi-ygc@umin.ac.jp
Abstract:
The Lsh/Ity/Bcg locus in the mouse genome regulates macrophage activation for antimicrobial activity against intracellular pathogens, and mouse Nramp1 (natural resistance-associated macrophage protein) gene was isolated as its candidate. The human NRAMP1 gene was subsequently isolated and its gene product was identified in macrophage/monocyte cells. Recently, a second Nramp gene, Nramp2, was found in mouse and human genomes. In the present study, we report the cloning and characterization of the human NRAMP2 gene, which is approximately 42 kb in length, containing 16 exons. The transcription start site was determined by 5'-RACE method, and the promoter was located between -246 bp to 145 bp in a region relative to the transcription start site, able to drive the luciferase reporter gene in HeLa cells. We also raised a polyclonal antibody against the glutathione S-transferase fusion protein containing the NH2-terminal 86 amino acids of human NRAMP2. The protein product of the human NRAMP2 gene is apparently present in human cultured cell lines as a 64 kDa protein recognized by this antibody, which is consistent with the molecular mass deduced from the human NRAMP2 cDNA.
Insights
Researchers cloned and characterized the human NRAMP2 gene, identifying its promoter region and confirming the protein
Area of Science:
- Molecular Biology
- Genetics
- Immunology
Background:
- The Lsh/Ity/Bcg locus regulates macrophage activation against intracellular pathogens.
- Mouse Nramp1 (natural resistance-associated macrophage protein) was identified as a candidate gene.
- Human NRAMP1 and a second gene, NRAMP2, have been identified in human genomes.
Purpose of the Study:
- To clone and characterize the human NRAMP2 gene.
- To identify the gene's promoter region and regulatory elements.
- To confirm the expression and molecular weight of the human NRAMP2 protein product.
Main Methods:
- Cloning and characterization of the human NRAMP2 gene (42 kb, 16 exons).
- Determination of the transcription start site using 5'-RACE.
- Promoter analysis using luciferase reporter gene assays in HeLa cells.
- Antibody generation against human NRAMP2 for protein detection.
Main Results:
- The human NRAMP2 gene spans approximately 42 kb and contains 16 exons.
- The promoter region was localized between -246 bp and 145 bp relative to the transcription start site.
- The human NRAMP2 protein product was detected as a 64 kDa protein in cultured cell lines.
Conclusions:
- The human NRAMP2 gene has been successfully cloned and characterized.
- Its promoter region demonstrates functional activity in driving gene expression.
- The presence and molecular weight of the human NRAMP2 protein are confirmed in human cells.
More Related Videos
10:05Hydrogel Nanoparticle Harvesting of Plasma or Urine for Detecting Low Abundance Proteins
Published on: August 8, 2014
11:21Forward Genetics Screens Using Macrophages to Identify Toxoplasma gondii Genes Important for Resistance to IFN-γ-Dependent Cell Autonomous Immunity
Published on: March 12, 2015
