[Agrobacterium tumefaciens mediated transformation of Aspergillus niger conidiospores and sequence analysis of T-DNA insertion site in conidiation-defective mutant]

Abstract

Objective: Development of a method of transforming Aspergillus niger conidiospores with Agrobacterium tumefaciens T-DNA system and using it as a tool for genome annotation via construction a T-DNA insertion mutant library of A. niger.

Methods: A. tumefaciens EHA105 carrying the binary vector pCAMBIA1301 was used to transform A. niger conidiospores under induction conditions. Hygromycin resistant transformants were screened on the selected medium. Sequence analysis was performed using inverse PCR method in stable mutants.

Results: In total, 193 hygromycin resistant stable transformants were obtained and the transformation rate was about 5.6 x 10(2) transformants /10(8) conidiospores. Obvious morphological changes were observed in some mutants and sequence analysis of T-DNA inserted site was performed in one conidiation-defective mutant. The result showed that the interrupted gene belonged to major facilitator superfamily (MFS).

Conclusion: Our results indicated that the transformation system of A. niger conidiospores set up in this study, combing with sequence analysis of T-DNA insertion site, might be an effective way for genome annotation in A. niger.