Construction of cDNA libraries from cultures of Armillaria mellea.
1. Conditions were established for growth of mycelial cultures of Armillaria mellea such that the production of its lysine-specific proteinase was maximized. Proteinase synthesis was confirmed by immunoprecipitation. 2. Mycelia grown under these same conditions were used as a source of RNA and this RNA was translatable in a wheat germ translation system to produce proteins with M(r) in the range < 10,000- > 90,000. 3. Double-stranded cDNA was prepared and was inserted into the EcoR1 site of lambda gt10 and lambda gt11 using an adaptor ligation strategy. Packaging of these materials yielded large cDNA libraries. The form lambda gt10 contained 2.9 x 10(6) pfu/ml with 70% recombinants whereas that from lambda gt11 contained 2.2 x 10(6) pfu/ml with 60% recombinants.
O'Connell
J
J
Department of Biochemistry, University College, Cork, Ireland.
Sheehan
H M
HM
Doonan
S
S
D000363:Agaricales / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D010582:Bacteriophage lambda / Q000235:genetics; D004247:DNA; D004274:DNA, Recombinant; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D005800:Genes, Fungal; D007163:Immunosorbent Techniques; D008970:Molecular Weight; D014176:Protein Biosynthesis; D012331:RNA, Fungal / Q000235:genetics / Q000302:isolation & purification
D004274:DNA, Recombinant; D012331:RNA, Fungal; D004247:DNA; D010450:Endopeptidases