Characterization and Cloning <i>of Pneumocystis carinii</i> Nucleic Acid

Abstract

Large numbers of Pneumocystis carinii (2 X 10(10) nuclei) were isolated and separated from the lungs of immunosuppressed rats by an enzymatic (collagenase, hyaluronidase and DNase) digestion procedure. The nucleic acid isolated from this P. carinii-enriched preparation was characterized by melting point analysis and RNA-sizing gels. The GC content of P. carinii DNA was approximately 33% while the rat DNA was 41.4%. In addition, RNA isolated from the P. carinii-enriched preparation showed unique ribosomal RNA bands of 3.4 kb and 1.8 kb as compared with uninfected rat lung ribosomal RNA which banded at 4.8 and 1.9 kb. Following isolation and fragmentation by sonication, the P. carinii DNA fragments were inserted into the vector, lambda gt-11. The resultant library contained 1.1 X 10(5) phage, of which 40-45% hybridized to P. carinii DNA but not to rat DNA.