Screening of Specific Probes from the DNA Library of Cryptococcus neoformans / 中华皮肤科杂志

ABSTRACT

To screen specific DNA probes by double hybridization from the constructed DNA li- brary of serotype A Cryptococcus neoformans.On the basis of the nucleotide sequence of vector pUC18, a pair of primers was synthesized.The insert fragments were amplified from the library on a PCR pro- cessor.The PCR products were spotted onto hybond-N~+ membranes.All inserts amplified from the ge- nomic library by PCR were screened by dot blot with 26 ~(32)P-labelled DNA from infectious agents for dif- ferentiation and from healthy persons.Twenty-eight candidate clones were obtained.The twenty-eight clone inserts got from low melting point agar were further characterized by dot blot with above 27 kinds of DNA for differentiation.Three specific DNA probes from the library of serotype A Cryptococcus neo- formans were obtained.Colony pCNIIA6 was serotype A-specific,which gave signals only with sertype A strain and did not cross hybridize with other DNAs.Colony pCNIIB5 was species-specific which gave signals only with DNA from Cryptococcus neoformans.Colony pCNIIIG1 was specific for var.neofor- mans,which gave signals only with serotype A and D strains.We can make a rapid diagnosis of Crypto- coccus neoformans infection at an early stage and distinguish the variants of C.neoformans and C.gattii using above specific probes.