Satyvaldy Jatayev

Drought poses a significant challenge to plant growth and productivity, particularly in arid regions like northern Kazakhstan. Dehydration-responsive element-binding (DREB) transcription factors play an important role in plant response to drought and other abiotic stresses. In Arabidopsis thaliana, the DREB subfamily consists of six groups, designated DREB1 to DREB6. Among these, DREB2 is primarily associated with drought and salinity tolerance. In the chickpea genome, two DREB genes, CaDREB2c and CaDREB2h, have been identified, exhibiting high sequence similarity to Arabidopsis DREB2 genes. We investigated the nucleotide sequences of CaDREB2c and CaDREB2h genes in several chickpea cultivars commonly grown in northern Kazakhstan. Interestingly, the CaDREB2h gene sequence was identical across all varieties and corresponded to the sequence deposited in the GenBank. However, the CaDREB2c gene sequence exhibited variations among the studied varieties, categorized into three groups: the first group (I), comprising 20 cultivars, contained a CaDREB2c gene sequence identical to the GenBank (Indian cultivar CDC Frontier). The second group (II), consisting of 4 cultivars, had a single synonymous substitution (T to C) compared to the deposited CaDREB2c gene sequence. The third group, encompassing 5 cultivars, displayed one synonymous substitution (C to T) and two non-synonymous substitutions (G to T and G to A). Furthermore, we assessed the gene expression patterns of CaDREB2c and CaDREB2h in different chickpea varieties under drought conditions. Chickpea cultivars 8 (III), 37 (I), 6 (III), and 43 (I) exhibited the highest drought resistance. Our analysis revealed a strong positive correlation between drought resistance and CaDREB2h gene expression under drought stress. Our findings suggest that the chickpea’s adaptive responses to water deprivation are associated with changes in CaDREB2 gene expression. To further elucidate the mechanisms underlying drought tolerance, we propose future research directions that will delve into the molecular interactions and downstream targets of CaDREB2 genes.

Salinity tolerance was studied in chickpea accessions from a germplasm collection and in cultivars from Kazakhstan. After NaCl treatment, significant differences were found between genotypes, which could be arranged into three groups. Those that performed poorest were found in group 1, comprising five ICC accessions with the lowest chlorophyll content, the highest leaf necrosis (LN), Na⁺ accumulation, malondialdehyde (MDA) content, and a low glutathione ratio GSH/GSSG. Two cultivars, Privo-1 and Tassay, representing group 2, were moderate in these traits, while the best performance was for group 3, containing two other cultivars, Krasnokutsky-123 and Looch, which were found to have mostly green plants and an exact opposite pattern of traits. Marker–trait association (MTA) between 6K DArT markers and four traits (LN, Na⁺, MDA, and GSH/GSSG) revealed the presence of four possible candidate genes in the chickpea genome that may be associated with the three groups. One gene, ATP-binding cassette, CaABCC6, was selected, and three haplotypes, A, D1, and D2, were identified in plants from the three groups. Two of the most salt-tolerant cultivars from group 3 were found to have haplotype D2 with a novel identified SNP. RT-qPCR analysis confirmed that this gene was strongly expressed after NaCl treatment in the parental- and breeding-line plants of haplotype D2. Mass spectrometry of seed proteins showed a higher accumulation of glutathione reductase and S-transferase, but not peroxidase, in the D2 haplotype. In conclusion, the CaABCC6 gene was hypothesized to be associated with a better response to oxidative stress via glutathione metabolism, while other candidate genes are likely involved in the control of chlorophyll content and Na⁺ accumulation.

Two genes, HvSAP8 and HvSAP16, encoding Zinc-finger proteins, were identified earlier as active in barley plants. Based on bioinformatics and sequencing analysis, six SNPs were found in the promoter regions of HvSAP8 and one in HvSAP16, among parents of two barley segregating populations, Granal × Baisheshek and Natali × Auksiniai-2. ASQ and Amplifluor markers were developed for HvSAP8 and HvSAP16, one SNP in each gene, and in each of two populations, showing simple Mendelian segregation. Plants of F₆ selected breeding lines and parents were evaluated in a soil-based drought screen, revealing differential expression of HvSAP8 and HvSAP16 corresponding with the stress. After almost doubling expression during the early stages of stress, HvSAP8 returned to pre-stress level or was strongly down-regulated in plants with Granal or Baisheshek genotypes, respectively. For HvSAP16 under drought conditions, a high expression level was followed by either a return to original levels or strong down-regulation in plants with Natali or Auksiniai-2 genotypes, respectively. Grain yield in the same breeding lines and parents grown under moderate drought was strongly associated with their HvSAP8 and HvSAP16 genotypes. Additionally, Granal and Natali genotypes with specific alleles at HvSAP8 and HvSAP16 were associated with improved performance under drought via higher 1000 grain weight and more shoots per plant, respectively.

Down-regulator associated protein, DrAp1, acts as a negative cofactor (NC2α) in a transcription repressor complex together with another subunit, down-regulator Dr1 (NC2β). In binding to promotors and regulating the initiation of transcription of various genes, DrAp1 plays a key role in plant transition to flowering and ultimately in seed production. TaDrAp1 and TaDrAp2 genes were identified, and their expression and genetic polymorphism were studied using bioinformatics, qPCR analyses, a 40K Single nucleotide polymorphism (SNP) microarray, and Amplifluor-like SNP genotyping in cultivars of bread wheat (Triticum aestivum L.) and breeding lines developed from a cross between spelt (T. spelta L.) and bread wheat. TaDrAp1 was highly expressed under non-stressed conditions, and at flowering, TaDrAp1 expression was negatively correlated with yield capacity. TaDrAp2 showed a consistently low level of mRNA production. Drought caused changes in the expression of both TaDrAp1 and TaDrAp2 genes in opposite directions, effectively increasing expression in lower yielding cultivars. The microarray 40K SNP assay and Amplifluor-like SNP marker, revealed clear scores and allele discriminations for TaDrAp1 and TaDrAp2 and TaRht-B1 genes. Alleles of two particular homeologs, TaDrAp1-B4 and TaDrAp2-B1, co-segregated with grain yield in nine selected breeding lines. This indicated an important regulatory role for both TaDrAp1 and TaDrAp2 genes in plant growth, ontogenesis, and drought tolerance in bread and spelt wheat.

Background: Chickpea is an important legume and is moderately tolerant to salinity stress during the growing season. However, the level and mechanisms for salinity tolerance can vary among accessions and cultivars. A large family of CaRab-GTP genes, previously identified in chickpea, is homologous to intracellular vesicle trafficking superfamily genes that play essential roles in response to salinity stress in plants. Results: To determine which of the gene family members are involved in the chickpea salt response, plants from six selected chickpea accessions (Genesis 836, Hattrick, ICC12726, Rupali, Slasher and Yubileiny) were exposed to salinity stress and expression profiles resolved for the major CaRab-GTP gene clades after 5, 9 and 15 days of salt exposure. Gene clade expression profiles (using degenerate primers targeting all members of each clade) were tested for their relationship to salinity tolerance measures, namely plant biomass and Na⁺ accumulation. Transcripts representing 11 out of the 13 CaRab clades could be detected by RT-PCR, but only six (CaRabA2, −B, −C, −D, −E and −H) could be quantified using qRT-PCR due to low expression levels or poor amplification efficiency of the degenerate primers for clades containing several gene members. Expression profiles of three gene clades, CaRabB, −D and −E, were very similar across all six chickpea accessions, showing a strongly coordinated network. Salt-induced enhancement of CaRabA2 expression at 15 days showed a very strong positive correlation (R² = 0.905) with Na⁺ accumulation in leaves. However, salinity tolerance estimated as relative plant biomass production compared to controls, did not correlate with Na⁺ accumulation in leaves, nor with expression profiles of any of the investigated CaRab-GTP genes. Conclusion: A coordinated network of CaRab-GTP genes, which are likely involved in intracellular trafficking, are important for the salinity stress response of chickpea plants.

Background: A family of genes designated as the Zinc finger A20/AN1 Transcription factors encoding stress-associated proteins (SAP) are well described in Arabidopsis and rice, and include 14 AtSAP and 18 OsSAP genes that are associated with variable tolerances to multiple abiotic stresses. The SAP gene family displays a great diversity in its structure and across different plant species. The aim of this study was to identify all HvSAP genes in barley (Hordeum vulgare L.), to analyse the expression of selected genes in response to salinity in barley leaves and develop SNP marker for HvSAP12 to evaluate the association between genotypes of barley plants and their grain yield in field trials. Results: In our study, 17 HvSAP genes were identified in barley, which were strongly homologous to rice genes. Five genes, HvSAP5, HvSAP6, HvSAP11, HvSAP12 and HvSAP15, were found to be highly expressed in leaves of barley plants in response to salt stress in hydroponics compared to controls, using both semi-quantitative RT-PCR and qPCR analyses. The Amplifluor-like SNP marker KATU-B30 was developed and used for HvSAP12 genotyping. A strong association (R² = 0.85) was found between KATU-B30 and grain yield production per plant of 50 F₃ breeding lines originating from the cross Granal × Baisheshek in field trials with drought and low to moderate salinity in Northern and Central Kazakhstan. Conclusions: A group of HvSAP genes, and HvSAP12 in particular, play an important role in the tolerance of barley plants to salinity and drought, and is associated with higher grain yield in field trials. Marker-assisted selection with SNP marker KATU-B30 can be applied in barley breeding to improve grain yield production under conditions of abiotic stress.

The relatively low efficiency of biolistic transformation and subsequent integration of multiple copies of the introduced gene/s significantly complicate the genetic modification of wheat (Triticum aestivum) and other plant species. One of the key factors contributing to the reproducibility of this method is the uniformity of the DNA/gold suspension, which is dependent on the coating procedure employed. It was also shown recently that the relative frequency of single copy transgene inserts could be increased through the use of nanogram quantities of the DNA during coating. A simplified DNA/gold coating method was developed to produce fertile transgenic plants, via microprojectile bombardment of callus cultures induced from immature embryos. In this method, polyethyleneglycol (PEG) and magnesium salt solutions were utilized in place of the spermidine and calcium chloride of the standard coating method, to precipitate the DNA onto gold microparticles. The prepared microparticles were used to generate transgenics from callus cultures of commercial bread wheat cv. Gladius resulting in an average transformation frequency of 9.9%. To increase the occurrence of low transgene copy number events, nanogram amounts of the minimal expression cassettes containing the gene of interest and the hpt gene were used for co-transformation. A total of 1538 transgenic wheat events were generated from 15,496 embryos across 19 independent experiments. The variation of single copy insert frequencies ranged from 16.1 to 73.5% in the transgenic wheat plants, which compares favourably to published results. The DNA/gold coating procedure presented here allows efficient, large scale transformation of wheat. The use of nanogram amounts of vector DNA improves the frequency of single copy transgene inserts in transgenic wheat plants.

KASP (KBioscience Competitive Allele Specific PCR) and Amplifluor (Amplification with fluorescence) SNP markers are two prominent technologies based upon a shared identical Allele-specific PCR platform. Amplifluor-like SNP and KASP analysis was carried out using published and own design of Universal probes (UPs) and Gene-specific primers (GSPs). Advantages of the Amplifluor-like system over KASP include the significantly lower costs and much greater flexibility in the adjustment and development of ‘self-designed’ dual fluorescently-labelled UPs and regular GSPs. The presented results include optimisation of ‘tail’ length in UPs and GSPs, protocol adjustment, and the use of various fluorophores in different qPCR instruments. The compatibility of the KASP Master-mix in both original and Amplifluor-like systems has been demonstrated in the presented results, proving their similar principles. Results of SNP scoring with rare alleles in addition to more frequently occurring alleles are shown. The Amplifluor-like system produces SNP genotyping results with a level of sensitivity and accuracy comparable to KASP but at a significantly cheaper cost and with much greater flexibility for UPs with self-designed GSPs.