State of the Art of Genetic Engineering in Potato: From the First Report to Its Future Potential

Potato (Solanum tuberosum L.) is a crop of world importance that produces tubers of high nutritional quality. It is considered one of the promising crops to overcome the challenges of poverty and hunger worldwide. However, it is exposed to different biotic and abiotic stresses that can cause significant losses in production. Thus, potato is a candidate of special relevance for improvements through conventional breeding and biotechnology. Since conventional breeding is time-consuming and challenging, genetic engineering provides the opportunity to introduce/switch-off genes of interest without altering the allelic combination that characterize successful commercial cultivars or to induce targeted sequence modifications by New Breeding Techniques. There is a variety of methods for potato improvement via genetic transformation. Most of them incorporate genes of interest into the nuclear genome; nevertheless, the development of plastid transformation protocols broadened the available approaches for potato breeding. Although all methods have their advantages and disadvantages, Agrobacterium-mediated transformation is the most used approach. Alternative methods such as particle bombardment, protoplast transfection with polyethylene glycol and microinjection are also effective. Independently of the DNA delivery approach, critical steps for a successful transformation are a rapid and efficient regeneration protocol and a selection system. Several critical factors affect the transformation efficiency: vector type, insert size, Agrobacterium strain, explant type, composition of the subculture media, selective agent, among others. Moreover, transient or stable transformation, constitutive or inducible promoters, antibiotic/herbicide resistance or marker-free strategies can be considered. Although great efforts have been made to optimize all the parameters, potato transformation protocols are highly genotype-dependent. Genome editing technologies provide promising tools in genetic engineering allowing precise modification of targeted sequences. Interestingly, transient expression of genome editing components in potato protoplasts was reported to generate edited plants without the integration of any foreign DNA, which is a valuable aspect from both a scientific and a regulatory perspective. In this review, current challenges and opportunities concerning potato genetic engineering strategies developed to date are discussed. We describe their critical parameters and constrains, and the potential application of the available tools for functional analyses or biotechnological purposes. Public concerns and safety issues are also addressed.

The interactome of CLUH reveals its association to SPAG5 and its co-translational proximity to mitochondrial proteins

Background Mitochondria require thousands of proteins to fulfill their essential function in energy production and other fundamental biological processes. These proteins are mostly encoded by the nuclear genome, translated in the cytoplasm before being imported into the organelle. RNA binding proteins (RBPs) are central players in the regulation of this process by affecting mRNA translation, stability, or localization. CLUH is an RBP recognizing specifically mRNAs coding for mitochondrial proteins, but its precise molecular function and interacting partners remain undiscovered in mammals. Results Here we reveal for the first time CLUH interactome in mammalian cells. Using both co-IP and BioID proximity-labeling approaches, we identify novel molecular partners interacting stably or transiently with CLUH in HCT116 cells and mouse embryonic stem cells. We reveal stable RNA-independent interactions of CLUH with itself and with SPAG5 in cytosolic granular structures. More importantly, we uncover an unexpected proximity of CLUH to mitochondrial proteins and their cognate mRNAs in the cytosol. We show that this interaction occurs during the process of active translation and is dependent on CLUH TPR domain. Conclusions Overall, through the analysis of CLUH interactome, our study sheds a new light on CLUH molecular function by revealing new partners and by highlighting its link to the translation and subcellular localization of some mRNAs coding for mitochondrial proteins.

Coordinated Translocation of Presequence-Containing Precursor Proteins Across Two Mitochondrial Membranes: Knowns and Unknowns of How TOM and TIM23 Complexes Cooperate With Each Other

The vast majority of mitochondrial proteins are encoded in the nuclear genome and synthesized on cytosolic ribosomes as precursor proteins with specific mitochondrial targeting signals. Mitochondrial targeting signals are very diverse, however, about 70% of mitochondrial proteins carry cleavable, N-terminal extensions called presequences. These amphipathic helices with one positively charged and one hydrophobic surface target proteins to the mitochondrial matrix with the help of the TOM and TIM23 complexes in the outer and inner membranes, respectively. Translocation of proteins across the two mitochondrial membranes does not take place independently of each other. Rather, in the intermembrane space, where the two complexes meet, components of the TOM and TIM23 complexes form an intricate network of protein–protein interactions that mediates initially transfer of presequences and then of the entire precursor proteins from the outer to the inner mitochondrial membrane. In this Mini Review, we summarize our current understanding of how the TOM and TIM23 complexes cooperate with each other and highlight some of the future challenges and unresolved questions in the field.

Challenges in Medicinal and Aromatic Plants DNA Barcoding—Lessons from the Lamiaceae

The potential value of DNA barcoding for the identification of medicinal plants and authentication of traded plant materials has been widely recognized; however, a number of challenges remain before DNA methods are fully accepted as an essential quality control method by industry and regulatory authorities. The successes and limitations of conventional DNA barcoding are considered in relation to important members of the Lamiaceae. The mint family (Lamiaceae) contains over one thousand species recorded as having a medicinal use, with many more exploited in food and cosmetics for their aromatic properties. The family is characterized by a diversity of secondary products, most notably the essential oils (EOs) produced in external glandular structures on the aerial parts of the plant that typify well-known plants of the basil (Ocimum), lavender (Lavandula), mint (Mentha), thyme (Thymus), sage (Salvia) and related genera. This complex, species-rich family includes widely cultivated commercial hybrids and endangered wild-harvested traditional medicines, and examples of potential toxic adulterants within the family are explored in detail. The opportunities provided by next generation sequencing technologies to whole plastome barcoding and nuclear genome sequencing are also discussed with relevant examples.

Genome structure and diversity among Cynanchum wilfordii accessions

Background Cynanchum wilfordii (Cw) and Cynanchum auriculatum (Ca) have long been used in traditional medicine and as functional food in Korea and China, respectively. They have diverse medicinal functions, and many studies have been conducted, including pharmaceutical efficiency and metabolites. Especially, Cw is regarded as the most famous medicinal herb in Korea due to its menopausal symptoms relieving effect. Despite the high demand for Cw in the market, both species are cultivated using wild resources with rare genomic information. Results We collected 160 Cw germplasm from local areas of Korea and analyzed their morphological diversity. Five Cw and one Ca of them, which were morphologically diverse, were sequenced, and nuclear ribosomal DNA (nrDNA) and complete plastid genome (plastome) sequences were assembled and annotated. We investigated the genomic characteristics of Cw as well as the genetic diversity of plastomes and nrDNA of Cw and Ca. The Cw haploid nuclear genome was approximately 178 Mbp. Karyotyping revealed the juxtaposition of 45S and 5S nrDNA on one of 11 chromosomes. Plastome sequences revealed 1226 interspecies polymorphisms and 11 Cw intraspecies polymorphisms. The 160 Cw accessions were grouped into 21 haplotypes based on seven plastome markers and into 108 haplotypes based on seven nuclear markers. Nuclear genotypes did not coincide with plastome haplotypes that reflect the frequent natural outcrossing events. Conclusions Cw germplasm had a huge morphological diversity, and their wide range of genetic diversity was revealed through the investigation with 14 molecular markers. The morphological and genomic diversity, chromosome structure, and genome size provide fundamental genomic information for breeding of undomesticated Cw plants.

Karyotype and plasmotype characteristics in wheat-rye hybrids with the different structural organization of the nuclear genome

Aim. Genome structure analysis and plasmotype identification in wheat-rye hybrids of various types (triti- cale, secalotriticum) and ploidy level. Mcth0ds. Cytological and molecular-genetic analysis. Rcsults. The karyotype and plasmotype analysis was carried out in 11 stable lines of secondary recombinant hexaploid triticale with the introgression of D-genome chromosomes of the wheat (A/B/DRR, 2n = 6x = 42), 14 stable and highly productive secalotriticum lines of F6–16 generations (Secalotriticum, S/RRAABB, 2n = 6x = 42), 9 stable lines of tetraploid triticale (A/BRR, 2n = 4x = 28). By means of differential chromosome staining, the chromosomal composition of the experimental material was characterized and the intergenomic substitution and translocation of chromosomes were detected. The PCR-RFLP analysis of the 18S/5S mitochondrial (mt) repeat and the ndhH-region of chloroplast DNA showed that these organ- elle DNA regions are in the homoplasmic state and belong to rye-type cytoplasm in secalotriticum lines and wheat-type cytoplasm in tetraploid and secondary recombinant hexaploid triticale lines. C0nclusi0ns. Cytological and molecular genetic analysis revealed significant genetic diversity of the created gene pool of wheat-rye hybrids by nuclear-cytoplasmic structure. The synthesized linear material of wheat-rye hybrids may be used in cytogenetic research and practical breeding.

Biological features and cultivation of sesame (a review)

Sesame is an ancient oilseed crop grown throughout the arid and subarid climates. Sesame oil has a good taste and is appreciated for its beneficial properties. The genus Sesamum L. (Pedaliaceae Lindl.) includes up to 38 species. The only cultivated species is S. indicum L. (2n = 26). It is grown in areas located between 45° N and 45° S. South Africa is considered the homeland of S. indicum. Sesame seeds contain 50–63% of oil, 25–27% of protein, 20% of carbohydrates, and are sources of such micronutrients as copper, calcium, iron, magnesium, zinc, vitamins A, B1, C and E. Sesame seeds and oil are used not only in the food industry but also in medicine. Sesame meal is a good animal feed. Recently, thanks to the deciphering of the sesame nuclear genome and the development of various genomic resources, including molecular markers for qualitative and quantitative characters, molecular breeding has become possible for such agronomic traits as high oil content and its composition, drought resistance, resistance to waterlogging, disease resistance, and high yield. The world production of sesame seeds according to FAO estimates for 2019 was about 7 million tons. The largest cropping areas are located in Africa, India, and China. In Russia, sesame can be grown in southern regions: Krasnodar and Stavropol Territories, Rostov and Astrakhan Provinces. Breeding work on sesame was carried out at the All-Union Research Institute of Oil Crops, where cultivars resistant to bacterial blight and Fusarium wilt were developed. Currently, sesame breeding programs are absent in Russia. There are over 25,000 sesame accessions in the world’s genebanks, including about 1,500 accessions in the VIR collection.

Insights from a comprehensive study of Trypanosoma cruzi: A new mitochondrial clade restricted to North and Central America and genetic structure of TcI in the region

More than 100 years since the first description of Chagas Disease and with over 29,000 new cases annually due to vector transmission (in 2010), American Trypanosomiasis remains a Neglected Tropical Disease (NTD). This study presents the most comprehensive Trypanosoma cruzi sampling in terms of geographic locations and triatomine species analyzed to date and includes both nuclear and mitochondrial genomes. This addresses the gap of information from North and Central America. We incorporate new and previously published DNA sequence data from two mitochondrial genes, Cytochrome oxidase II (COII) and NADH dehydrogenase subunit 1 (ND1). These T. cruzi samples were collected over a broad geographic range including 111 parasite DNA samples extracted from triatomines newly collected across NA and CA, all of which were infected with T. cruzi in their natural environment. In addition, we present parasite reduced representation (Restriction site Associated DNA markers, RAD-tag) genomic nuclear data combined with the mitochondrial gene sequences for a subset of the triatomines (27 specimens) collected from Guatemala and El Salvador. Our mitochondrial phylogenetic reconstruction revealed two of the major mitochondrial lineages circulating across North and Central America, as well as the first ever mitochondrial data for TcBat from a triatomine collected in Central America. Our data also show that within mtTcIII, North and Central America represent an independent, distinct clade from South America, named here as mtTcIIINA-CA, geographically restricted to North and Central America. Lastly, the most frequent lineage detected across North and Central America, mtTcI, was also an independent, distinct clade from South America, noted as mtTcINA-CA. Furthermore, nuclear genome data based on Single Nucleotide Polymorphism (SNP) showed genetic structure of lineage TcI from specimens collected in Guatemala and El Salvador supporting the hypothesis that genetic diversity at a local scale has a geographical component. Our multiscale analysis contributes to the understanding of the independent and distinct evolution of T. cruzi lineages in North and Central America regions.