Mitochondrial Sequence Diversity Among Alloplasmic and Euplasmic Triticum Species

Four mitochondrial genomes of Triticum species were sequenced and annotated: 1) (lo) durum mitochondrial genome, which is an alloplasmic line with Triticum longissimum(SS) cytoplasm and T. turgidum (AABB) nucleus, 2) T. longissimum, 3) T. turgidum and 4) T. tauschii (DD). Comparison showed major differences in atp6, nad9, nad6, rps19-p, cob and cox2 genes among all four species. Additionally, species-specific ORFs were also identified. A single nucleotide polymorphism search within known genes showed that the alloplasmic line differs from the two parental lines by six nucleotides in the cox3, mttB, rps2, rps4 and rps13 genes. We were able to recognize mitochondrial heteroplasmy based on single nucleotide variation (SNV) and regions of high SNV density within a given species. Structural differences between T. turgidum, (lo) durum and T. longissimum mitochondrial genomes were observed; however, conserved gene blocks and gene pairs among these species were identified. Three possible recombination events in gene blocks I, V and VI were recognized. We observed differences in the alloplasmic line, compared to its parental lines in: sequence, predicted genes, single nucleotide polymorphism (SNP) and genome structure. These facts support the hypothesis of the accelerated evolution of the mitochondrial genome when transferred into alien nuclear background. We also found that major gene changes recognized here appear to be common among Triticum species. Based on sequence assembly, we report full mitochondrial sequence of T. turgidum. We recognized 40 SNP differences compared to the T. aestivum mitochondrial genome, where 5 SNPs were found in known mitochondrial genes: rps1, rps2, cox3 and ccmFN. The T. longissimum and T. tauschii share highly similar genomes in structure and content, different only at the level of SNVs. A method to establish phylogenetic relationships based on mitochondrial genome sequence is proposed using differences in reference assembly with a common mitochondrial backbone sequence. We confirm conservation of the mitochondrial gene content within Triticum species. These results create background to explore the role of mitochondrial genes in conditioning nuclear cytoplasmic incompatibility in a wide range of Triticum alloplasmic lines and also deepen our understanding of evolutionary relationships that exist in the Triticum genus.US Army Research Offic

Structure and function of class III pistil-specific extensin-like protein in interspecific reproductive barriers

Abstract Background The transmitting tissue of the style is the pathway for pollen tube growth to the ovules and has components that function in recognizing and discriminating appropriate pollen genotypes. In Nicotiana tabacum, the class III pistil extensin-like (PELPIII) arabinogalactan protein is essential for the inhibition of N. obtusifolia pollen tube growth. The transmitting tissue-specific (TTS) arabinogalactan protein amino acid sequence and expression pattern is similar to PELPIII, but it facilitates self-pollinated N. tabacum. The TTS and PELPIII arabinogalactan protein can be divided into the less conserved N-terminal (NTD) and the more conserved C-terminal (CTD) domains. This research tested whether the NTD is the key domain in determining PELPIII function in the inhibition of interspecific pollen tube growth. Three variant PELPIII gene constructs were produced where the PELPIII NTD was exchanged with the TTS NTD and a single amino acid change (cysteine to alanine) was introduced into the PELPIII NTD. The PELPIII variants of N. tabacum were tested for activity by measuring the inhibition N. obtusifolia pollen tube growth by using them to complement a 3’UTR RNAi transgenic line with reduced PELPIII mRNA. Results The RNAi N. tabacum line had reduced PELPIII mRNA accumulation and reduced inhibition of N. obtusifolia pollen tube growth, but had no effect on self-pollen tube growth or pollen tube growth of 12 other Nicotiana species. The NTD of PELPIII with either the PELPIII or TTS CTDs complemented the loss PELPIII activity in the RNAi transgenic line as measured by inhibition of N. obtusifolia pollen tube growth. The TTS NTD with the PELPIII CTD and a variant PELPIII with a cysteine to alanine mutation in its NTD failed to complement the loss of PELPIII activity and did not inhibit N. obtusifolia pollen tube growth. Conclusion The NTD is a key determinant in PELPIII’s function in regulating interspecific pollen tube growth and is a first step toward understanding the mechanism of how PELPIII NTD regulates pollen tube growth

ALICE 実験 PHOS 検出器による運動量凍結温度測定法

Primers used during cDNA synthesis, 5′ and 3′ RACE and gene specific product amplification. (DOCX 20 kb

Additional file 3: Table S3. of Polymorphism and structure of style–specific arabinogalactan proteins as determinants of pollen tube growth in Nicotiana

A. Summary of the disorder disposition prediction for stylar AGP and NtPRP. (DOCX 53 kb