Organelle Comparative Genome Analysis Reveals Novel Alloplasmic Male Sterility with orf112 in Brassica oleracea L.

B. oleracea Ogura CMS is an alloplasmic male-sterile line introduced from radish by interspecific hybridization and protoplast fusion. The introduction of alien cytoplasm resulted in many undesirable traits, which affected the yield of hybrids. Therefore, it is necessary to identify the composition and reduce the content of alien cytoplasm in B. oleracea Ogura CMS. In the present study, we sequenced, assembled, and compared the organelle genomes of Ogura CMS cabbage and its maintainer line. The chloroplast genome of Ogura-type cabbage was completely derived from normal-type cabbage, whereas the mitochondrial genome was recombined from normal-type cabbage and Ogura-type radish. Nine unique regions derived from radish were identified in the mitochondrial genome of Ogura-type cabbage, and the total length of these nine regions was 35,618 bp, accounting for 13.84% of the mitochondrial genome. Using 32 alloplasmic markers designed according to the sequences of these nine regions, one novel sterile source with less alien cytoplasm was discovered among 305 materials and named Bel CMS. The size of the alien cytoplasm in Bel CMS was 21,587 bp, accounting for 8.93% of its mtDNA, which was much less than that in Ogura CMS. Most importantly, the sterility gene orf138 was replaced by orf112, which had a 78-bp deletion, in Bel CMS. Interestingly, Bel CMS cabbage also maintained 100% sterility, although orf112 had 26 fewer amino acids than orf138. Field phenotypic observation showed that Bel CMS was an excellent sterile source with stable 100% sterility and no withered buds at the early flowering stage, which could replace Ogura CMS in cabbage heterosis utilization.

Effect of Alien Cytoplasm on Combining Ability for Earliness and Seed Yield in Sunflower under Irrigation and Drought Stress

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Sequencing and annotation of the chloroplast DNAs and identification of polymorphisms distinguishing normal male-fertile and male-sterile cytoplasms of onion

Male-sterile (S) cytoplasm of onion is an alien cytoplasm introgressed into onion in antiquity and is widely used for hybrid seed production. Owing to the biennial generation time of onion, classical crossing takes at least 4 years to classify cytoplasms as S or normal (N) male-fertile. Molecular markers in the organellar DNAs that distinguish N and S cytoplasms are useful to reduce the time required to classify onion cytoplasms. In this research, we completed next-generation sequencing of the chloroplast DNAs of N- and S-cytoplasmic onions; we assembled and annotated the genomes in addition to identifying polymorphisms that distinguish these cytoplasms. The sizes (153 538 and 153 355 base pairs) and GC contents (36.8%) were very similar for the chloroplast DNAs of N and S cytoplasms, respectively, as expected given their close phylogenetic relationship. The size difference was primarily due to small indels in intergenic regions and a deletion in the accD gene of N-cytoplasmic onion. The structures of the onion chloroplast DNAs were similar to those of most land plants with large and small single copy regions separated by inverted repeats. Twenty-eight single nucleotide polymorphisms, two polymorphic restriction-enzyme sites, and one indel distributed across 20 chloroplast genes in the large and small single copy regions were selected and validated using diverse onion populations previously classified as N or S cytoplasmic using restriction fragment length polymorphisms. Although cytoplasmic male sterility is likely associated with the mitochondrial DNA, maternal transmission of the mitochondrial and chloroplast DNAs allows for polymorphisms in either genome to be useful for classifying onion cytoplasms to aid the development of hybrid onion cultivars.

Investigations of IsogenomicAlloplasmics of Flue-Cured Tobacco Nicotianatabacum cv. Wislica

Cytoplasms of fifteen wild Nicotianae and a male sterile cytoplasm (cms) from N. tabacum were bred into the genome of a standard Polish flue-cured cultivar Wislica. The sixteen iso-genomic alloplasmics were compared for expression of male sterility and for some traits related to field performance. In cmsN. bigelovii, N. debneyi, N. exigua, N. megalosiphonor N. suaveolens stamens were absent, in cmsN. tabacum, N. glauca, N. goodspeedii or N. undulata stamens were absent or rudimentary, in cmsN. eastii, N. glutinosa or N. plumbaginifoliastamen tissue became petaloid, in those from N. knightiana and N. raimondiithey were morphologically normal. Female organ morphology was changed in cmsN. goodspeedii, N. occidentalis, N. exigua, N. debneyior N. bigelovii, seed set was reduced only in cmsN. occidentalis. Plant height in most alloplasmics was similar to that of the fertile variety but was strongly depressed by the cytoplasms of N. glutinosa, N. eastiiand N. plumbaginifolia. Leaf area was positively affected by N. amplexicaulis, N. bigeloviiand N. undulatacytoplasms, unaffected by N. suaveolens, N. tabacumand N. glaucacytoplasms and negatively affected by the remaining cytoplasms mostly because of the narrowing of the leaf blade. Cured leaf yields from cmsN. bigelovii were higher than those of Wislica. Leaf yields from cmsN. amplexicaulis, N. suaveolens, N. glauca or N. tabacum were not affected by the alien cytoplasm, whereas those from the remaining alloplasmics were depressed to different extents. Increased incidence of PVY and white spots (mostly attributable to Cercospora sp.) were observed on many of the alloplasmics especially on cmsN. exigua and N. suaveolens(PVY) and increased white spots only on cmsN. bigelovii, N. exigua, N. occidentalis and N. undulata.

Interspecific androgenetic restoration of rosy barb using cadaveric sperm

Interspecific androgenetic rosy barb (Puntius conchonius) was generated using its cadaveric (-20 °C) or fresh sperm to activate nuclear genome inactivated oocytes of gray tiger barb (Puntius tetrazona). UV irradiation was used to inactivate nuclear genome of tiger barb oocytes. Thermal shock restored diploidy of rosy barb in the oocytes of tiger barb. Survival of androgenotes was 14% or 7% when fresh or cadaveric sperm was used. The diploid or haploid nuclear genome of rosy barb, individually or jointly with that of tiger barb, regulated the time sequence of embryonic development in an alien cytoplasm of tiger barb oocytes. Androgenetic males (Y2Y2) attained sexual maturity earlier and had significantly higher gonadosomatic index and sperm concentration, albeit suffering a slight decrease in fertilizing ability. Conversely, androgenetic females (X2X2) suffered extended interspawning period, reduced fecundity, and poor hatchability of their progenies. These results are discussed with respect to their significance for conservation biology.Key words: nuclear genome inactivation, tiger barb, cadaveric sperm, rosy barb, interspecific androgenotes, Tc1 transposon.

Interactions between the scs and Vi genes in alloplasmic durum wheat

Two nuclear genes, vitality (Vi) on an A- or B-genome chromosome and species cytoplasm specific (scs) on a 1DL telosome from Triticum aestivum L. or a telosome from Aegilops uniaristata Vis. (un telosome), improved compatibility between the nucleus of Triticum turgidum L. var. durum and the cytoplasm of Ae. longissima S. &M. or Ae. uniaristata. To study interactions between Vi and scs and to determine the chromosomal location of Vi, 29-chromosome fertile plants were crossed with 13 D-genome disomic-substitution (d-sub) lines [except 5D(5A)] of 'Langdon' durum. F1 and backcross progenies were examined for meiotic chromosome number and pairing, fertility, and plant vigor. In 11 crosses, Vi restored seed viability but produced double-monosomics (d-monos) with greatly reduced growth and vigor. In contrast, crosses involving 1D(1A) and 1D(1B) d-sub lines produced d-monos with normal vigor and anthesis but nonfunctional pollen. A backcross of 1D + 1A d-mono F1 and 1D(1A) d-sub lines produced 11 male steriles; 3 had 13 II + 1 II 1D + 1 I 1A, 2 had 13 II + 2 I, 1 had 13 II + 1 II 1D(1A), and 5 were not examined. Crosses of 1D + 1A d-mono F1 with control durum, lo durum (with 1DL), and un durum (with un telosome) lines produced 16 male-sterile d-monos and 14 fertiles with 14 II + 1 I 1D, showing that 15-chromosome female gametes transmitted monosomes 1A and 1D. However, BC2F1's from 1D + 1B d-mono × fertile line with un telosome included 20 male-sterile d-monos, 6 fertile triple monosomics (13 II + 1 I 1D + 1 I 1B + t I un telosome), and 1 fertile plant with a 1B/1D translocation. Unlike d-mono 1A + 1D, d-mono 1B + 1D did not transmit 15-chromosome female gametes with monosomes 1D and 1B. Additional backcrosses also indicated that homozygous scs caused male sterility in 1D(1A) and 1D(1B) d-subs and that the procedure used was not suitable for the chromosomal location of Vi.Key words: alien cytoplasm, nucleocytoplasmic interactions, 1B/1D translocation, aneuploidy, cytoplasmic male sterility.