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Molecular Cloning of a Neurospora crassa Carotenoid Biosynthetic Gene (Albino-3) Regulated by Blue Light and the Products of the White Collar Genes

1989; Taylor & Francis; Volume: 9; Issue: 3 Linguagem: Inglês

10.1128/mcb.9.3.1271-1276.1989

1098-5549

Mary Anne Nelson, Giorgio Morelli, Alessandra Carattoli, Nicoletta Romano, Giuseppe Macino,

Algal biology and biofuel production

The albino-3 (al-3) gene of Neurospora crassa, which probably encodes the carotenoid biosynthetic enzyme geranylgeranyl pyrophosphate synthetase, was cloned. The N. crassa triple mutant al-3 qa-2 aro-9 was transformed to qa-2+ with mixtures of plasmids bearing N. crassa DNA inserts, and the transformants were screened for the al-3+ phenotype. One al-3+ qa-2+ transformant (AL3-1) was examined in detail and shown to contain intact vector sequences integrated into the N. crassa genome. The vector and some flanking sequences were recovered from AL3-1 after restriction, ligation, and selection of chloramphenicol-resistant transformants of Escherichia coli. The flanking sequences were subsequently used to detect the al-3-containing plasmid in the mixture of about 1,800 plasmids. Restriction fragment length polymorphism mapping was carried out to confirm the identity of the cloned fragment. The level of the al-3 mRNA was shown to be increased 15-fold in light-induced (compared with that in dark-grown) wild-type mycelia. The light-dependent increase in al-3 mRNA levels was not observed in presumed regulatory mutant (white collar) strains.