The mitochondrial DNA of diplonemid and kinetoplastid protists is known for its suite of bizarre features, including the presence of concatenated circular molecules, extensive trans-splicing and various forms of RNA editing. Here we report on the existence of another remarkable characteristic: hyper-inflated DNA content. We estimated the total amount of mitochondrial DNA in four kinetoplastid species (Trypanosoma brucei, Trypanoplasma borreli, Cryptobia helicis, and Perkinsela sp.) and the diplonemid Diplonema papillatum. Staining with 4',6-diamidino-2-phenylindole and RedDot1 followed by color deconvolution and quantification revealed massive inflation in the total amount of DNA in their organelles. This was further confirmed by electron microscopy. The most extreme case is the ∼260 Mbp of DNA in the mitochondrion of Diplonema, which greatly exceeds that in its nucleus; this is, to our knowledge, the largest amount of DNA described in any organelle. Perkinsela sp. has a total mitochondrial DNA content ~6.6× greater than its nuclear genome. This mass of DNA occupies most of the volume of the Perkinsela cell, despite the fact that it contains only six protein-coding genes. Why so much DNA? We propose that these bloated mitochondrial DNAs accumulated by a ratchet-like process. Despite their excessive nature, the synthesis and maintenance of these mtDNAs must incur a relatively low cost, considering that diplonemids are one of the most ubiquitous and speciose protist groups in the ocean. © 2018 IUBMB Life, 70(12):1267-1274, 2018.

• MeSH
• Euglenozoa genetics   MeSH
• Phylogeny MeSH
• Kinetoplastida genetics   MeSH
• DNA, Mitochondrial genetics   isolation & purification   ultrastructure   MeSH
• Mitochondria genetics   MeSH
• Trans-Splicing genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Aneuploidies in embryos conceived by assisted reproduction techniques can originate before meiosis, during meiosis and after fertilization. To evaluate the risk of aneuploidy from the maternal meiosis, it is possible to analyze the first polar body (PB1) and second polar body (PB2). The paternal pre-meiotic and meiotic error rate can be evaluated by sperm analysis. Postfertilization errors can only be analyzed by embryo biopsy. Methods: Molecular cytogenetic methods for analysis of gametes are based on fluorescence in situ hybridization (FISH) and fluorescence microscopy. New options include use of peptide nucleic acid probes (PNA-FISH) and primed in situ synthesis (PRINS) for detection of chromosome specific sequences. The standard 2-dimensional image analysis can be complemented by 3- dimensional analysis using Z-stacking and deconvolution (3D-FISH) for more precise detection of FISH signals in interphase nuclei that are not flat after fixation (sperm, cells from blastocysts). Results: We have tested use of FISH, PNA-FISH, PRINS and 3D-FISH for detection of aneuploidy in sperm as well as FISH and PNA-FISH for analysis of first and second polar bodies and oocytes. We have also tested FISH, PNA-FISH and 3D-FISH for analysis of blastomeres as well as FISH and 3D-FISH for reanalysis of embryos in blastocyst stage after abnormal results of preimplantation screening of aneuploidy. Conclusions: For optimal evaluation of risk of aneuploidy after conception it is possible to combine the prefertilization sperm analysis with polar body and blastomere biopsy after fertilization. This complex evaluation allows optimal choice of reproductive medicine methods and early and efficient prevention of embryonal/fetal aneuploidies.

• Publication type
• Abstracts MeSH

• MeSH
• Cell Biology MeSH
• Molecular Biology MeSH
• Publication type
• Monograph MeSH

• Conspectus
• Biochemie. Molekulární biologie. Biofyzika
• NML Fields
• biologie
• cytologie, klinická cytologie