Characterization of a novel plasmid-like element inNeurospora crassaderived mostly from the mitochondrial DNA

Abstract

We have identified a plasmid-like element within mitochondria of Neurospora crassa strain stp-B1. It is derived from the EcoRI-4 and EcoRI-6 regions of the mitochondrial DNA, and an additional 124 bp DNA segment of unknown origin. The plasmid DNA consists of an oligomeric series of circular molecules of monomer length 2.2 kbp. The abundance of the plasmid suggests its autonomous replication and the presence of an efficient origin of replication. An unusually large number of palindromes capable of forming secondary structures are present in the plasmid. Such a palindrome, located near sequences reminiscent of mammalian and fungal mtDNA origins of replication, may define the replication origin of the plasmid. This putative origin might also represent the replication origin of the wild-type mtDNA.

Recombination by sequence repeats with formation of suppressive or residual mitochondrial DNA in Neurospora.

“…SA). Similar secondary structures were found in E35, pSNM, and stp-B1, the only Neurospora mtDNA mutants studied earlier (13,20,21) for which the DNA sequence of the recombination junction is known (Fig. 5 B and C).…”

“…It represents a plasmid-like element derived from the mtDNA by excision and subsequent amplification of a contiguous DNA from EcoRI-4/6 wild-type mtDNA. Its detailed characterization including the recombination junction and entire nucleotide sequence was determined (13). In contrast to the above findings, AA9, abn-2r, and ER-3r displayed the pattern of restriction fragments expected for the wild type, suggesting the absence of recombinant mtDNA molecules.…”

“…The Neurospora strains were obtained from the Fungal Genetics Stock Center (FGSC) (Kansas City, KS). The wild-type strain was RL3-8A (FGSC 2218) and the spontaneous mitochondrial mutant strains were abn-J (FGSC 1448), abn-2 (FGSC 1458) (12), and stp-BI (FGSC 1574) (13). All other strains were generated in our laboratory.…”

Large scale solubilization of coal and bioconversion to utilizable energy. Quarterly report, October 1--December 31, 1996

“…The plasmids pYEleu-2 and pstp 2.2 and the genomic and cDNA libraries of Neurospora and PCR machine are available in my laboratory. All methods of molecular cloning, transformation, and identification and characterization of transformants will be as practiced in my laboratory (Schablik et al, 1982;Almasan and Mishra, 1988;1990; or as described previously (Hinnen et al, 1978; Maniatis et al, 1989;Yadav and Mishra, 1994;Feher and Mishra, 1994).…”

Large scale solubilization of coal and bioconversion to utilizable energy. Fifth quarterly technical report, January 1, 1995--March 31, 1995

“…All methods of molecular cloning, transformation, and identification and characterization of transformants will be as practiced in my laboratory (Schablik et al, 1982;Almasan and Mishra, 1988;1990; 1991 1 or as described previously (Hinnen et al, 1978;Maniatis et al, 1989;Yadav and Mishra, 1994;Feher and Mishra, 1994).…”