[Immunological screening of Toxoplasma tachyzoite cDNA expression libraries with serum from infected rats].
OBJECTIVE
To screen and identify the potential candidates for the development of toxoplasmosis vaccine.


METHODS
Rats were infected with Toxoplasma gondii (T. gondii) RH strain and their sera were used as a probe to screen T. gondii tachyzoite cDNA expression libraries. The positive clones were analyzed by PCR amplification and DNA sequencing.


RESULTS
Thirteen positive clones were obtained from about 4 x 10(5) phage plagues after three rounds of screening. The size of the inserts ranged from 0.45 kb to 2.4 kb. A BLAST search of all available sequence databases using the partial sequences from four positive clones (L1, L2, L4, L5) showed that the sequence of L2 clone was identical with T. gondii P24 major antigen gene (TgP24). Clone L4 had a high homology with Saccharum officinarum pyruvate orthophosphate dikinase. There is no significant hit of any sequences to clone L1, suggesting that L1 could be a novel gene(GenBank accession number AY180109), named T.g-R1, which encodes a non-transmembrane protein with 134 amino acid open reading frame. PROSCAN analysis of the T.g-R1 amino acid sequence showed that this gene product contains two protein kinase C phosphorylation site, two casein kinase II phosphorylation site, one N-myristoylation site and one microbodies C-terminal targeting signal. Clone L5 was a small partial fragment.


CONCLUSIONS
The identification of positive clones provides a possible way for the development of toxoplasmosis vaccine.
Jiang
Li-ping
LP
Department of Parasitology, Xiangya School of Medicine, Central South University, Changsha 410078.
Wu
Xiang
X
Cai
Li-ting
LT
Wang
Dan-jing
DJ
Shu
Heng-ping
HP
D000595:Amino Acid Sequence; D000818:Animals; D000953:Antigens, Protozoan / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D005260:Female; D015723:Gene Library; D008969:Molecular Sequence Data; D051381:Rats; D017207:Rats, Sprague-Dawley; D014122:Toxoplasma / Q000235:genetics* / Q000276:immunology
D000953:Antigens, Protozoan; D018076:DNA, Complementary