Genotype identification and diversity analysis in Kiwifruit (Actinidia spp.) using RAPD markers

Kiwifruit (Actinidia spp.) is a significant plantation crop belonging to family Actinidiaceae, having deciduous, dioecious, and scrambling vines with chromosome number 2n=58. Dioecy in kiwifruit forms the basis for several breeding programs. The present study was carried out for diversity analysis in kiwifruit genotypes using RAPD markers. 7 kiwifruit genotypes (2 males viz. Allision & Tomuri and 5 females viz. Hayward, Bruno, Allision, Monty & Abbott) were analysed for molecular polymorphism using 94 RAPD primers, out of which 23 primers amplified the genomic DNA in all the genotypes. RAPD data was analysed using NTSYS-pc software and dendrogram construction was done using UPGMA method. Two separate clusters of male and female genotypes were formed. Similarity matrix indices showed maximum similarity between Tomuri (M) and Allision (M) with a similarity coefficient of 0.719 while Abbott (F) and Allision (M) were found to have least similarity having a similarity coefficient of 0.521. Four RAPD primers amplified unique amplicons in Monty, Hayward, Bruno, Allision (M) and Abbott and two primers amplified unique amplicons in Allision (M) & Tomuri (M) along with the male and female plants of Allision genotype respectively. Therefore, these primers can help in distinguishing the genotypes of kiwifruit and can also be validated as putative markers for the sex identification in kiwifruit.

Efficiency of SSR and PawS markers for evaluation of genetic polymorphism among red clover (Trifolium pratense L.) cultivars

Background. Identification of crop varieties is presently one of the most important aspects due a significant annual increase in the number of newly developed cultivars. Application of molecular markers makes it possible to identify cultivars and secure protection of plant breeders’ rights. Marker techniques based on SSR loci and PawS markers were evaluated for their efficiency in revealing the DNA polymorphism in Russian red clover cultivars, and the research results are presented in this publication.Materials and Methods. The total genome DNA was extracted by a modified SDS method from 30 seedlings per each cultivar. Nine simple sequence repeats (SSR) and 4 PawS markers were used for genotyping. The basic genetic diversity parameters were measured and analyzed using the software resources GelAnalyzer 2010а, MStools v.3, and Statistica 7.0.Results and conclusion. The mean level of intervarietal DNA polymorphism in red clover was 38.6%. Cultivar-specific amplicons were obtained for 4 accessions (cvs. ‘Trifon’, ‘Topaz’, ‘Trio’ and ‘Mars’) with SSR loci RCS1307 and RCS3095. These loci were found appropriate for identification and certification of such cultivars. The tested PawS markers (individually and in combinations) proved non-informative for the analysis of intervarietal DNA polymorphism in red clover. The only primer pair PawS5+PawS16 generated reproducible PCR products, but unique amplicons were absent in the DNA profiles. The data obtained in this study may be helpful for further identification and certification of Russian red clover cultivars and promising breeding materials.

Characterization of Pigeonpea Genotypes for Waterlogging Tolerance Based on Morpho-physiological and Molecular Traits

Aim: To screen and characterize pigeonpea genotypes using morpho-physiological, biochemical and molecular traits. Study Design: The field trial was conducted using the Randomized Block Design (RBD) while Completely Randomized Design was used for laboratory and pot screening experiments. Place and Duration of Study: The research trial was conducted at Laboratory and Pulse Research Farm, Model Bhitti, BAU, Sabour, which lies between 25°15’40” N latitude to 87°2’42” E longitude and 46 meters above sea level. Study was undertaken between July, 2018 to March, 2019. Methodology: Sixty pigeonpea genotypes were screened for submergence tolerance at seed stage in the laboratory. Based on results of laboratory screening, 40 genotypes with sufficient genotypic variability for waterlogging tolerance were further taken for seedling stage screening at field, finally 20 genotypes were taken to pots for waterlogging tolerance evaluation and characterization on the basis of morpho-physiological, biochemical and molecular traits. According to pot results, six contrasting genotypes were considered for RAPD primers amplification. Results: The three levels of sieving of genotypes fetched results directing the opportunity of particular genotypes to be sown inlow land areas. Character like seed colour varied from brown, dark brown to black, which showed significant relationship with level of tolerance. Significant (p=0.01) higher germination with less reduction due to waterlogging stress was observed in genotypes such as ICP-11809, ICPL-20098, NDA-1 and ICP-5028. Maximum survival percent was found in ICP-5028 (62.28%) while least survival percent was observed in ICP-7035 (10.98%). At the field stage, genotypes such as ICP-5028, ICPL-990985, ICPL-20238 were best performing genotypes. SPAD Chlorophyll Content results exhibited significant reduction among the susceptible genotypes. However, there was least reduction among tolerant genotypes such as LRG-30, Mal-9, Pusa-992 and ICP-5028.Genotypes namely: Manak, Pusa-991 and Pusa-992 faced hastened senescence under waterlogging condition as compared to ICP-5028, Mal-15, Mal-9, LRG-30. Molecular evaluation results of six genotypes chosen across screening showed that ICP-7035 and Manak were clubed together in one cluster. Nevertheless, ICP-5028 and Mal-9 were grouped in another cluster of Dendrogram, constructed using Jaccard similarity coefficient. In the present investigation two unique amplicons were amplified by primers OPA-13 and OPC-01. OPA-13 amplified unique band was linked with susceptible genotypes of size ~1240 bp while the unique amplicon given by OPC-01 was of ~980 bp size linked with tolerant genotypes. Conclusion: Available waterlogging tolerance in pigeonpea gene pool to some extent can provide source for breeding waterlogging tolerant cultivars. Physiological and genetic approach involving efficient screening techniques and evaluation of breeding material/lines under targeted environment for the traits linked to tolerance is likely to lead to the identification of specific component traits and high yielding varieties with improved stress tolerance.