Genetic diversity and population structure of sweet orange [Citrus sinensis (L.) Osbeck] germplasm of India revealed by SSR and InDel markers

Sweet orange (Citrus sinensis (L.) Osbeck) is an important commercial citrus fruit crop, cultivated in India and across the world. In India most of the cultivated sweet orange species were introduced varieties. In this study, we used two molecular markers, SSR and InDels, to understand the genetic diversity and population structure of seventy-two sweet orange genotypes. Genetic parameters consisted of a total number of alleles, a number of polymorphic alleles (effective alleles); genetic diversity (G.D.), expected heterozygosity (He), and the polymorphic information content (PIC) were calculated based on molecular data. Two dendrograms were constructed based on the InDels and SSR. In both the cases, they formed three major clusters showing various degrees of variations with respect to members of the clusters. Population structure analysis revealed the presence of two distinct subpopulations. Therefore, in order to address various challenges and develop sweet orange varieties with desirable traits, there is a need to broaden the genetic base of sweet orange through the intensive collection in the northeastern region. These results of intraspecific genetic variability of the collections will dictate the path for the sweet orange breeding and conservation programs in India.

Mapping of dwarfing QTL of Ari1327, a semi-dwarf mutant of upland cotton

Background Upland Cotton (Gossypium hirsutum L.) has few cotton varieties suitable for mechanical harvesting. The plant height of the cultivar is one of the key features that need to modify. Hence, this study was planned to locate the QTL for plant height in a 60Co γ treated upland cotton semi-dwarf mutant Ari1327. Results Interestingly, bulk segregant analysis (BSA) and genotyping by sequencing (GBS) methods exhibited that candidate QTL was co-located in the region of 5.80–9.66 Mb at D01 chromosome in two F2 populations. Using three InDel markers to genotype a population of 1241 individuals confirmed that the offspring’s phenotype is consistent with the genotype. Comparative analysis of RNA-seq between the mutant and wild variety exhibited that Gh_D01G0592 was identified as the source of dwarfness from 200 genes. In addition, it was also revealed that the appropriate use of partial separation markers in QTL mapping can escalate linkage information. Conclusions Overwhelmingly, the results will provide the basis to reveal the function of candidate genes and the utilization of excellent dwarf genetic resources in the future.

Genetic Structure and Forensic Feature of 38 X-Chromosome InDels in the Henan Han Chinese Population

Insertion/deletion (InDel) polymorphisms, as ideal forensic markers, show useful characteristics of both SNPs and STRs, such as low mutation rate, short amplicon size and general applicability of genotyping platform, and have been used in human identification, population genetics and biogeographic research in recent years. X-chromosome genetic markers are significant in population genetic studies and indispensable complements in some complex forensic cases. However, the population genetic studies of X-chromosome InDel polymorphisms (X-InDels) still need to be explored. In this study, the forensic utility of a novel panel including 38 X-InDel markers was evaluated in a sample of Han population from Henan province in China. It is observed that the heterozygosities ranged from 0.0054 to 0.6133, and the combined discrimination power was 1–9.18 × 10−17 for males and 1–7.22 × 10−12 for females respectively. The mean exclusion chance in trios and duos were 0.999999319 and 0.999802969 respectively. Multiple biostatistics methods, such as principal component analysis, genetic distances analysis, phylogenetic reconstruction, and structure analysis was used to reveal the genetic relationships among the studied Henan Han group and other 26 reference groups from 1,000 Genomes Project. As expected, the Henan Han population was clustered with East Asian populations, and the most intimate genetic relationships existed in three Han Chinese populations from Henan, Beijing and South China, and showed significant differences compared with other continental groups. These results confirmed the suitability of the 38 X-InDel markers both in individual identification and parentage testing in Han Chinese population, and simultaneously showed the potential application in population genetics.

Developing functional markers for vitamin E biosynthesis in oil palm

Vitamin E is essential for human health and plays positive roles in anti-oxidation. Previously, we detected large variation in vitamin E content among 161 oil palm accessions. In this study, twenty oil palm accessions with distinct variation in vitamin E contents (171.30 to 1 258.50 ppm) were selected for genetic variation analysis and developing functional markers associated with vitamin E contents. Thirty-seven homologous genes in oil palm belonging to vitamin E biosynthesis pathway were identified via BLASTP analysis, the lengths of which ranged from 426 to 25 717 bp (average 7 089 bp). Multiplex PCR sequencing for the 37 genes found 1 703 SNPs and 85 indels among the 20 oil palm accessions, with 226 SNPs locating in the coding regions. Clustering analysis for these polymorphic loci showed that the 20 oil palm accessions could be divided into five groups. Among these groups, group I included eight oil palm accessions whose vitamin E content (mean value: 893.50 ppm) was far higher than other groups (mean value 256.29 to 532.94 ppm). Correlation analysis between the markers and vitamin E traits showed that 134 SNP and 7 indel markers were significantly (p < 0.05) related with total vitamin E content. Among these functional markers, the indel EgTMT-1-24 was highly correlated with variation in vitamin E content, especially tocotrienol content. Our study identified a number of candidate function associated markers and provided clues for further research into molecular breeding for high vitamin E content oil palm.

Marker-Assisted Evaluation of Two Powdery Mildew Resistance Candidate Genes in Korean Cucumber Inbred Lines

Two genes, CsLRR-RPK2 (CsGy5G015660) and CsaMLO8 (Csa5G623470), have been considered as powdery mildew (PM) resistance genes in cucumbers. In this study, we evaluated the involvement of the alleles of these two genes in PM resistance in 100 commercial Korean cucumber inbred lines. To achieve this, we developed cleaved amplified polymorphic sequences (CAPS) and InDel markers from CsLRR-RPK2 and CsaMLO8. Genotyping analysis indicated that the CsLRR-RPK2-CAPS marker showed a stronger correlation with the PM-resistant phenotype, with an 84% consistency compared to the CsaMLO8-InDel marker. The use of the CsaMLO8-InDel marker showed a 70% consistency between phenotype and genotype results. It was proposed that the CsLRR-RPK2-CAPS marker successfully eliminated PM-susceptible inbred lines, since both genotype and phenotype results were 100% identical. Furthermore, the present study revealed that the introduction of one of these alleles is probably enough to confer PM resistance in cucumbers. However, seven PM-resistant inbred lines harbored either CsaMLO8 or CsLRR-RPK2 alleles, indicating that there is another PM-resistant resource(s) besides CsaMLO8- and CsLRR-RPK2–originated resistance in the commercial Korean inbred lines. Our results provide reliable evidence confirming two PM-resistant candidate genes for the detection of PM resistance resources in cucumber inbred lines.

Geographical diversity of Helicobacter pylori strains circulating in the European Part of the Russian Federation

The aim of the study was to identify INDEL markers and study geographical origin of regional H. pylori strains circulating in the European part of the Russian Federation. The study included 56 strains of H. pylori isolated in three regions of the Russian Federation: Saint Petersburg, Astrakhan and Rostov Regions. Genomic DNA was isolated using a set of Probe NA (DNA Technology, Russia), according to the manufacturer’s instructions. Detection of INDEL markers hp5605, hp6405, hp340, hp1390, hp3660 was performed using PCR. Clustering of the identified INDEL genotypes and building a phylogenetic tree were performed using the BioNumerics 7.6 and GrapeTree software packages. 21 strains from the GenBank database with known geographical origin were used as reference strains. In 20 strains from Saint Petersburg, 13 individual genotypes were identified, while 17 strains belong to the European cluster (hpEurope), 2 strains belong to the hspEAsia cluster and one strain belongs to the hspWAfrica cluster. The most common genotype identified in the European cluster includes six strains from Saint Petersburg and two strains from the GenBank database. For further differentiation of these strains, the VNTR typing method was used, which allowed identifying eight individual genotypes in eight strains. Fifty-six studied russian strains are represented by thirty individual genotypes, which reflects the high heterogeneity of strains circulating in the European part of the Russian Federation. The most frequent genotype is represented by two hpEurope strains, one strain from the Astrakhan region, as well as 5 and 6 strains from the Rostov Region and Saint Petersburg, respectively. The vast majority of Russian strains (52/56) belong to the hpEurope population, while two strains from Saint Petersburg are included in the hspEAsia population, and one strain from Saint Petersburg and the Astrakhan Region is included in the hspWAfrica population. Total, 77 H. pylori strains are represented by 37 individual genotypes with a high diversity index (DI = 0.95), which allows us to consider the proposed INDEL typing method as an independent method for genotyping H. pylori strains. Taking into consideration the complexity of the problem of accurately determining the geographical origin of H. pylori strains, the proposed simple and convenient method of INDEL typing of H. pylori strains, based on an available PCR method becomes very relevant and allows us to conduct an adequate primary analysis of the geographical origin of Russian H. pylori strains.

Transcriptomic analysis of salt tolerance-associated genes and diversity analysis using indel markers in yardlong bean (Vigna unguiculata ssp. sesquipedialis)

Background High salinity is a devastating abiotic stresses for crops. To understand the molecular basis of salinity stress in yardlong bean (Vigna unguiculata ssp. sesquipedalis), and to develop robust markers for improving this trait in germplasm, whole transcriptome RNA sequencing (RNA-seq) was conducted to compare the salt-tolerant variety Suzi 41 and salt-sensitive variety Sujiang 1419 under normal and salt stress conditions. Results Compared with controls, 417 differentially expressed genes (DEGs) were identified under exposure to high salinity, including 42 up- and 11 down-regulated DEGs in salt-tolerant Suzi 41 and 186 up- and 197 down-regulated genes in salt-sensitive Sujiang 1419, validated by qRT-PCR. DEGs were enriched in “Glycolysis/Gluconeogenesis” (ko00010), “Cutin, suberine and wax biosynthesis” (ko00073), and “phenylpropanoid biosynthesis” (ko00940) in Sujiang 1419, although “cysteine/methionine metabolism” (ko00270) was the only pathway significantly enriched in salt-tolerant Suzi 41. Notably, AP2/ERF, LR48, WRKY, and bHLH family transcription factors (TFs) were up-regulated under high salt conditions. Genetic diversity analysis of 84 yardlong bean accessions using 26 InDel markers developed here could distinguish salt-tolerant and salt-sensitive varieties. Conclusions These findings show a limited set of DEGs, primarily TFs, respond to salinity stress in V. unguiculata, and that these InDels associated with salt-inducible loci are reliable for diversity analysis.