Application of SCoT markers for accessing of genetic diversity of gramineous forage grass species

Using the SCoT marker system, 8 varieties of cereal grasses belonging to 5 species were analyzed: Festuca pratensis, Lolium perenne, Lolium multiflorum, Festuca rubra, Festulolium. Of the 10 tested SCoT markers, 7 informative markers were selected that reveal interspecies genetic polymorphism. According to the results of the analysis, DNA profiles characteristic of each studied species were obtained, and primers allowing to detect intervarietal differences for subsequent identification and molecular genetic passportization were selected.

Cryopreservation of Arecanut (Areca catechu L.) Embryogenic Calli by V Cryo-plate Method

Aims: Arecanut, a perennial palm species of Arecaceae family, has huge commercial value, and is grown mainly for its masticatory nuts. The ever-increasing demand for uniform quality plantlets from growers necessitates putting in place In vitro mass multiplication and other crop improvement programmes. The present study was carried out to standardize the procedure for cryopreservation of embryogenic calli of arecanut, derived from immature inflorescence cultures, by vitrification based cryo-plate technique. Study Design: Completely randomized design (CRD) with three replications. Place and Duration of Study: ICAR-Central Plantation Crops Research Institute, Kerala, India during 2019. Methodology: The embryogenic calli were precultured in Eeuwen's Y3 basal medium supplemented with sucrose (0.2, 0.3 and 0.4 M) for three days. Explants were affixed on cryo-plates and later dehydrated using plant vitrification solution 3 (PVS3) for 30 min. Cryoplates were inserted in cryovials and cryopreserved. Explants with no cryostorage served as control. Explants were rewarmed quickly in a water bath (40ºC) for 2 min and treated with unloading solution and cultured on recovery medium. Results: The results showed 8-10 % recovery of embryogenic calli that resulted in normal plantlet production. The clonal fidelity studies, using Start Codon Targeted (SCoT) marker, showed no variation of cryopreserved calli in comparison to the original calli. Conclusion: This preliminary study demonstrated the successful use of vitrification (V) cryo-plate technique in cryopreservation of embryogenic calli of arecanut. With better recovery percentage, the optimal concentration of sucrose in the preculture medium was found to be 0.3 M. Desiccation in PVS3 solution for 30 min had no adverse effect.

SCoT Markers Analysis of Genetic Diversity of Polygonatum, A Medicinal and Edible Plant

Background: As a traditional Chinese medicine, Polygonatum has been demonstrated to have immunomodulatory, antibacterial, anti-inflammatory, anti-aging, anti-cancer, hypoglycemic and other pharmacological effects. However, the germplasm resources of Polygonatum have been destroyed in recent years and the research on its genetic diversity is extremely scarce. In this study, the genetic diversity of 28 Polygonatum germplasms from 11 different provinces in China was evaluated by Start codon targeted (SCoT) marker. Results: A total of 365 bands were generated by 15 SCoT primers, of which 355 were polymorphic, with a high polymorphism of 97.3%. And the genetic similarity coefficient is between 0.59 and 0.75, indicating a high genetic diversity. UPGMA (Unweighted Pair Group Method with Arithmetic Mean) dendrogram, PCoA (Principal Coordinate Analysis) and Structure analysis have similar results in grouping Polygonatum germplasm and they are all divided into two populations. We found that there was a certain correlation between the genetic distance and geographical distance of Polygonatum germplasm. By analyzing other valid genetic diversity parameters (Na, Ne, H, I), it is clarified that Polygonatum has abundant alleles and abundant genetic diversity among populations. Conclusions: This study suggests that Polygonatum germplasm resources from different provenances have rich genetic diversity. The SCoT molecular marker is a valuable marker system and can be used for genetic analysis of Polygonatum resources. This will be helpful to further study the preservation and genetic improvement of Polygonatum germplasm.

Genetic variability among three Egyptian isolates of Heterorhabditis indica using a new marker technique (SCoT)

Background Entomopathogenic nematodes (EPNs) are a group of nematode families, have the ability to search for their hosts, and are considered as promising biological control candidates for insect pests, providing protection to non-target organisms and the environment. Results This study was conducted to isolate indigenous EPN isolates from Egyptian agricultural soils for further use in biological control programs and study their genetic polymorphism among the previously isolated isolates under accession no. MH553167 and MK300683 and the new isolate (MH496627), using the start codon targeted (SCoT) marker. One out of 15 soil samples obtained from a banana cultivated field was positive for the presence of EPNs, using the Galleria baiting method. Morphological analysis and sequencing of the internal transcribed spacer (ITS) region suggested that the isolate obtained belongs to Heterorhabditis indica. The sequence of the ITS was submitted to the National Center for Biotechnology Information (NCBI) and registered under accession no. MH496627. Ten SCoT primers were used in the study; the polymorphic bands were 68 out of 76 with 89% as polymorphism percentage. The highest numbers of bands were 10 bands generated by SCoT 1 and SCoT 18 while SCoT 48 and SCoT 60 recorded the lowest band number (5 bands). Conclusions The present study is considered as a preliminary study to demonstrate the effectiveness of the SCoT marker for the first time in assessing genetic relationships in EPNs.

Identification of Local Citrus Breeds Using the Start-Codon Targeted Polymorphism Technique Combined with Clonal Sequencing

This study explored the genetic diversity of 35 citrus accessions using start-codon targeted (SCoT) markers. Total 15 primers were used to amplify products ranging in length from 100 to 2000 bp. A total of 133 fragments were amplified and found that 126 (95%) were polymorphic. The genetic similarity coefficients among the 35 accessions ranged from 0.67 to 1.00, indicating that the SCoT markers could reveal high genetic diversity in the citrus germplasm. A cluster analysis showed that local citrus breeds of Sihui city fell into one cluster, possibly reflecting the geographical distribution of the tested samples. The sizesof the fragments of eight local citrus cultivars amplified by one of the primers ranged from 951 to 1001 bp, and the similitude was 95.17%high. Both single-base mutations, insertions and deletions were identified among the fragments and comparison with asequence database suggested that the amplified region was part of a ribosomal protein-coding sequence. Using Scot marks combined with clonal sequencing, each of the test samples had one or more mutation sites that could be used as markers to differentiate from the other seven test samples. Thus, the resolution of the genetic diversity among the local citrus breeds revealed by the SCoT technique was enhanced by the subsequent sequencing analysis of specific fragments. The experimental results also provide evidence that the relationship between CitrusnobilisL our. ‘gonggan’ and C. hanianaHort. ex Tseng ‘Sihuihanggan’ is that between parent and offspring hybrids and not between bud-mutation strains. The SCoT marker is a targeted gene molecular marker, based on the characteristic bands of primers, it can be used as a marker to isolate genes of local citrus varieties and also for investigating mutational hotspots of the segene.© 2021Friends Science Publishers