Cloning a novel mouse Gabarapl2 cDNA and its characterization
Abstract
Aim: To clone a novel mouse GABAA-receptor-associated protein like 2 (Gabarapl2) gene, and to analysis its primary function.
Methods: With the aid of computer, the human GABARAPL2 cDNA was used as information probe to search mouse EST database of GenBank for mouse homolog. A series of overlapping EST were found and assembled into an EST contig using Genetics Computer Group (GCG) ASSEMBLY program. The existence of the gene was then identified by experiment. Northern blotting was performed to hybridize [alpha-32P]dATP labeled probe with mRNA of 11 different mouse tissues that had been transferred to the nylon membrane.
Results: The novel gene was deposited in GenBank under Accession No AF190644. Its cDNA contained an intact open reading frame and a canonical polyadenylation signal AATAAA followed by polyA. The deduced protein was completely identical to that of human GABARAPL2, and was termed Gabarapl2 by Mouse Gene Nomenclature Committee. The putative protein of Gabarapl2 has a calculated molecular weight of 13 700 and an isoelectric point of 8.56. It was also predicted to contain two protein kinase C phosphorylation sites and one tyrosine kinase phosphorylation site. Northern hybridization showed that Gabarapl2 was expressed as a single 1.35 kb transcript, with high levels in brain, thymus, lung, heart, kidney, and liver, and low in pancreas, testis, small intestine, colon, and stomach.
Conclusion: A novel mouse Gabarapl2 gene was cloned and identified.
Keywords:
Methods: With the aid of computer, the human GABARAPL2 cDNA was used as information probe to search mouse EST database of GenBank for mouse homolog. A series of overlapping EST were found and assembled into an EST contig using Genetics Computer Group (GCG) ASSEMBLY program. The existence of the gene was then identified by experiment. Northern blotting was performed to hybridize [alpha-32P]dATP labeled probe with mRNA of 11 different mouse tissues that had been transferred to the nylon membrane.
Results: The novel gene was deposited in GenBank under Accession No AF190644. Its cDNA contained an intact open reading frame and a canonical polyadenylation signal AATAAA followed by polyA. The deduced protein was completely identical to that of human GABARAPL2, and was termed Gabarapl2 by Mouse Gene Nomenclature Committee. The putative protein of Gabarapl2 has a calculated molecular weight of 13 700 and an isoelectric point of 8.56. It was also predicted to contain two protein kinase C phosphorylation sites and one tyrosine kinase phosphorylation site. Northern hybridization showed that Gabarapl2 was expressed as a single 1.35 kb transcript, with high levels in brain, thymus, lung, heart, kidney, and liver, and low in pancreas, testis, small intestine, colon, and stomach.
Conclusion: A novel mouse Gabarapl2 gene was cloned and identified.