Molecular characterization reveals chlorosis-corrected CMS (Brassica oxyrrhina) B. juncea cybrid has recombinant mitochondrial genome involving male sterility inducing orf108-atpA gene

Abstract

Alloplasmic Brassica juncea lines with B. oxyrrhina
cytoplasm show stable cytoplasmic male sterility (CMS)
but display chlorosis due to incompatible interaction
between the plastid and the nuclear genomes. Normal, green
CMS cybrids were developed through somatic hybridization
between chlorotic CMS line and B. juncea. This study was
aimed at characterization of the mitochondrial genome
constitution of two chlorosis-corrected B. juncea cybrids.
Comparative RFLP analysis of parents and cybrids with
mitochondrial gene specific probes revealed B. oxyrrhina
specific pattern in cybrids for atp4, ccmfn2, nad4L and
cox3 but atp1 and orf108 gave novel pattern indicating
recombinant mitochondria. RT-PCR showed cotranscription of orf108 and atp1 in the cybrids. PCR analysis
with mitochondrial genome specific primers revealed that
the cybrids had mostly B. oxyrrhina like mitochondrial
genome. The two cybrids showed identical RFLP and PCR
pattern suggesting that they are derived from the same
event. Interestingly, RFLP pattern indicated two copies of
orf108-atp1 in the cybrid. One of the copies was derived
from intermolecular recombination between B. juncea and
B. oxyrrhina whereas the second copy resulted from
intramolecular recombination between a pair of short repeat
regions. The results show that orf108-atp1 region is
recombinationally active and could be involved in causing
male sterility