ABSTRACT
A total of 17,439 mature miRNAs (~ 21 nt) earlier generated through RNA seq in the pomegranate were used for in silico analysis. After complexity reduction, a total of 1922 representative mature miRNAs were selected and used as query sequences against pomegranate genome to retrieve 2540 homologous contigs with flanking regions (~ 800). By using pre-miRNA prediction web server, a total of 1028 true contigs harbouring pri-miRNAs encoding 1162 pre-miRNAs were identified. Survey of these sequences for SSRs yielded a total of 1358 and 238 SSRs specific to pri-miRNA and pre-miRNAs, respectively. Of these, primer pairs were designed for 897 pri-miRNA and 168 pre-miRNA SSRs. In pri-miRNA sequences, hexa-nucleotides repeats were found to be most abundant (44.18%) followed by mono- (18.41%) and di-nucleotide (17.01%), which is also observed in pre-miRNA sequences. Further, a set of 51 randomly selected pre-miRNA-SSRs was examined for marker polymorphism. The experimental validation of these markers on eight pomegranate genotypes demonstrated 92.15% polymorphism. Utility of these functional markers was confirmed via examination of genetic diversity of 18 pomegranate genotypes using 15 miRNA-SSRs. Further, potential application of miRNA-SSRs for discovery of trait specific candidate genes was showed by validating 51 mature miRNA against publically available 2047 EST sequences of pomegranate by target and network analysis. In summary, the current study offers novel functional molecular markers for pomegranate genetic improvement.
ABSTRACT
The present study investigates the genetic diversity and population structure among 42 diverse pomegranate genotypes using a set of twenty one class I hypervariable SSR markers (> 24 bp), which were reported earlier from the analysis of cv. Dabenzi genome. The study material comprised 16 indigenous and 13 exotic cultivars, and 13 wild accessions. A total of 66 alleles (Na) were detected with an average of 3.14 alleles per marker. The average values of polymorphic information content (PIC), observed heterozygosity (Ho) and Shannon's gene diversity index (I) were 0.44, 0.21 and 0.95, respectively suggesting moderate genetic diversity. The pairwise genetic distance ranged from 0.07 to 0.80 with a mean value of 0.53. Population structure analysis divided all the genotypes into four subpopulations (SP1, SP2, SP3 and SP4). Interestingly, the results of phylogenetic and principal component analyses coincided with the results of structure analysis and the grouping of genotypes followed the geographical origins. AMOVA revealed that 25% of the variation was attributed to differences among populations, whereas 75% within the subpopulations with significant F ST value 0.25 (p < 0.001), indicating a high level of genetic differentiations or low level of gene flow. Based on the F ST values, pomegranate genotypes belonging to SP4 (indigenous cultivars) followed by SP1 (exotic lines) exhibited higher gene diversity and genetic differentiations within and among populations. These genetic relationships based on SSR markers could be harnessed in future genetic improvement of pomegranate through informed hybridization programs.
ABSTRACT
This study reports genome wide characterization and development of first set of microsatellite markers through in silico analysis of eight sequenced Xanthomonas axonopodis pv. punicae strains available in the public database. SSR survey resulted in identification of ~ 4638 perfect SSRs, with mean marker frequency 901 SSRs/Mb and densitiy of 11,006 bp/Mb aross the eight genomes. Frequency distribution graphs revealed hexa-nucleotide repeats were more prominent fowllowed by tri-, tetra-, di- and penta-nucleotides in the analysed genomes. We desinged 2927 SSR primers that are specific to the strain LMG 859 and ePCR confirmed on seven other Xap genomes. This resulted in identification of 542 informative SSRs that are producing single amplicons, from which 66 primers were successfully validated through wet lab experiments on eight Xap isolates of pomegranate. Furthermore, utility of these SSRs were demostrated by analysing molecular diversity among 22 Xap isolates using 20 Xap_SSR primers. SSRs revealed moderate genetic diversity among Xap isolates (61%) and grouped 11 isolates that are repersenting six different states into one cluster. This proved the earlier evidence of wider spread of ST3 type Xap acoss India using Multi locus Sequence Typing (MLST) technique. In summary, Xap_SSR will serve as powerful genomics tools that would helps in monitoring of population dynamics, taxonomy, epidomology and quarantine aspects in bacterial blight pathogen through development of microsatellite based Multilocus Variable number of Tandem repeat analysis (MLVA) in future. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03209-z.
ABSTRACT
The alkaloids N-methylhydrasteine hydroxylactam and 1-methoxyberberine chloride were isolated from Corydalis longipes. Both alkaloids showed high efficacy individually (in concentration of 50-150 ppm) and also in a 1:1 mixture against spore germination of some fungi, viz. Alternaria alternata, A. brassicae, Curvularia maculans, Curvularia sp., Colletotrichum gloeosporioides, Colletotrichum sp., Helminthosporium speciferum, H. pennisetti, Helminthosporium sp., and Ustilago cynodontis. The antifungal effect of single compounds was dose-dependent. If the mutual ratio of the two components in the mixture was changed from 1:1 to a major content of any of the two compounds, the inhibitory effect on spore germination decreased.