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Molecular characterization and n use efficiency of LeAlaAT ‘Mekongga’ transgenic rice lines


Yulita DS, Purwoko BS, Sisharmini A, Apriana A, Santoso TJ, Trijatmiko KR, Sukma D

SUMMARY

Genetic engineering is one of the strategies for developing nitrogen (N)-use-efficient rice (Oryza sativa) varieties. One gene that plays an indirect role in N metabolism is alanine aminotransferase (AlaAT). It can efficiently increase N content and crop yield. In a previous study, the tomato AlaAT gene (LeAlaAT) was successfully isolated and introduced into ‘Mekongga’ rice. The present research was conducted during 2018 and 2019 at the Indonesian Center for Agricultural Biotechnology and Genetic Resources Research and Development (ICABIOGRAD), Bogor, Indonesia. The objectives of the present study were to perform the molecular characterization of LeAlaAT ‘Mekongga’ rice lines on the basis of the hpt marker gene, the direct PCR of the LeAlaAT fragment, and the phenotypic evaluation of the selected LeAlaAT T1 ‘Mekongga’ rice lines in response to different N fertilizer rates (0 kg ha−1 [control] and 60, 90, and 120 kg ha−1). This research involved three activities, namely (1) Southern blot analysis, (2) direct PCR, and (3) N use efficiency (NUE) test of ‘Mekongga’ transgenic lines. Southern blot analysis revealed that in T0 transgenic lines, the copy number of the hpt marker gene varied from 1 to 3. Direct PCR confirmed the presence of the AlaAT fragment in the T1 generation of five ‘Mekongga’ transgenic lines. The five transgenic lines showed high panicle number, biomass weight, shoot dry weight, and total grain weight under 120 kg ha−1 nitrogen. The high agronomical NUE of transgenic lines under 120 kg ha−1 N implied that the transgenic rice lines have the potential for efficient N use at a certain minimum level of N (120 kg ha−1 of nitrogen) and should be further evaluated at high N levels.

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Keyword: Ma Alanine aminotransferase, LeAlaAT transgenic rice lines, Mekongga, Southern blot, direct PCR, N use efficiency

DOI: https://doi.org/10.54910/sabrao2021.53.4.14

Assessment of tea plant (Camellia sinensis L.) accessions for pollen sources in natural crossing by using microsatellites

Azka NA, Taryono, Wulandari RA

SUMMARY

Tea (Camellia sinensis L. [O.] Kuntze) is a highly cross-pollinated and self-incompatible plant. Seeds can be harvested from specific individual mother plants in polyclonal tea gardens. Whether the pollen donor plays an important role in seed formation remains unclear. This study aimed to identify the male parents of 72 natural hybridized progenies (F1) from one female parent on the basis of a putative specific allele by using simplesequence repeat (SSR) markers and the exclusion-likelihood method with Cervus 3.0 software. The genetic material, which comprised seven accessions of C. sinensis L., was acquired from Assamica planted in the Kayulandak polyclonal seed garden of the Pagilaran tea plantation in Batang District, Central Java, Indonesia, and was studied during 2019 and 2020. The genotype PGL-15 was used as the female parent, whereas the six candidate genotypes PGL-10, GMB-9, GMB-7, TPS-93, GMB-11, and TRI 2025 were used as the male parents. In this study, 13 SSR loci were used to identify the male parents of the F1 progenies obtained through natural hybridization between one female and six male tea accessions. Results indicated that the exclusion-likelihood method, which correctly predicted 100% of the male parents, was more effective than the putative specific allele approach, which correctly predicted only 34.72% of the male parents in the 72 hybridized F1 progenies of tea plants.

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Keywords: Camellia sinensis L., natural pollination, SSR markers, paternity analysis, putative specific allele, exclusion-likelihood method

DOI: https://doi.org/10.54910/sabrao2021.53.4.10

Rice backcross population assessment for iron tolerance through phenotypic and genotypic analyses

Tam VT, vy LT, Huu NT, Ha PTT

SUMMARY

Iron toxicity has become a serious issue affecting rice (Oryza sativa L.) production in many irrigated lowland areas. The selection of Fe2+-tolerant rice cultivars under iron toxicity conditions and the identification of molecular markers are good approaches to obtaining tangible results. This study aimed to identify simple sequence repeat (SSR) markers that were associated with iron tolerance traits in a rice backcross population. A total of 117 seedlings from the backcross (BC3F2) of ‘OM6830’/‘AS996’//‘AS996’ were phenotyped at the 4-week-seedling stage at Ton Duc Thang University, Ho Chi Minh City, Vietnam. The rice population was screened in Yoshida nutrient medium supplemented with FeCl2 at a concentration of 150 mg L−1 under greenhouse conditions. Phenotypic analysis was conducted by scoring two parameters, namely, root length and leaf bronzing. Genotypic analysis was carried out on the BC3F2 population by using four markers, i.e., RM6, RM240, RM252, and RM451, for association analysis with iron tolerance. A total of 23 BC3F2 lines were selected on the basis of their higher tolerance (score 1) for Fe2+ compared with the tolerant parental line ‘AS996’. The markers RM6 and RM240 were highly polymorphic and identified different Fe2+-tolerant lines in the BC3F2 population. Among the BC3F3 progeny derived from the selected 23 BC3F2 lines, BC3F3-7 was identified as the most Fe2+-tolerant line. BC3F3-15 was also found to be Fe2+ tolerant. Both lines showed good development capability and provided high yields under stress conditions. These tolerant BC3F3 lines could be further screened with additional SSR markers in future breeding programs aiming to increase rice production in iron-contaminated areas of the Mekong Delta, Vietnam.

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Keywords: Backcrosses, iron toxicity, phenotypic and genotypic traits, screening, SSR markers, genetic analysis, Oryza sativa L.

DOI: https://doi.org/10.54910/sabrao2021.53.4.7

Analysis of the genetic diversity of Phalaenopsis orchids with single nucleotide polymorphisms and snap markers derived from the Pto gene

Sukma D, Elina J, Raynalta E, Aisyah SI, Aziz SA, Sudarsono, Chan MT

SUMMARY

The Pto gene is a plant gene that has been reported to be involved in resistance to bacterial pathogens. A partial genomic sequence corresponding to Pto (~449 bp) was isolated from 16 species and four hybrids of Phalaenopsis during 2017 at the Department of Agronomy and Horticulture, IPB University, Bogor, Indonesia. Multiple sequence analysis was performed to find putative single nucleotide polymorphisms (SNPs) and design the corresponding single nucleotide-amplified polymorphism (SNAP) markers, which were in turn used to estimate the genetic diversity of 25 Phalaenopsis species. In total, 20 SNPs, of which 14 were nonsynonymous, were identified from the partial Pto sequences. Eighteen SNAP primers were then developed based on these 14 nonsynonymous and four synonymous SNPs. Validation results showed that 15 SNAP primers showed a polymorphism information content exceeding 0.3, suggesting the existence of more than two alleles for this locus. Upon their use, the SNAP markers described 86% of all interspecies variability. The Pto 52, Pto 349, Pto 229, and Pto 380 SNAP markers were very informative in the determination of genetic diversity. Notably, the existence of these nonsynonymous SNPs implied the possibility of functional changes within the amino acid sequence of the putative PTO protein. Thus, the resulting differences in the activity of the PTO protein may be used to breed tolerance to pathogen infection. Further work may be required to establish a functional link between tolerance to pathogens and the presence of Pto-SNAP markers in Phalaenopsis properly.

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Keywords: Phalaenopsis, moth orchid, diversity, single nucleotide polymorphism, Pto, bacterial resistance

DOI: https://doi.org/10.54910/sabrao2021.53.4.6

Genotype by environment interaction analysis of wheat (Triticum aestivum L.) grain yield under rainfed conditions in Zambia

Tembo B

SUMMARY

Understanding genotype by environment interaction (GEI) is important for crop improvement because it aids in the recommendation of cultivars and the identification of appropriate production environments. The objective of this study was to determine the magnitude of GEI for the grain yield of wheat grown under rain-fed conditions in Zambia by using the additive main effects and multiplicative interaction (AMMI) model. The study was conducted in 2015/16 at Mutanda Research Station, Mt. Makulu Research Station and Golden Valley Agricultural Research Trust (GART) in Chibombo. During2016/17, the experiment was performed at Mpongwe, Mt. Makulu Research Station and GART Chibombo, Zambia. Fifty-five rain-fed wheat genotypes were evaluated for grain yield in a 5 × 11 alpha lattice design with two replications. Results revealed the presence of significant variation in yield across genotypes, environments, and GEI indicating the differential performance of genotypes across environments. The variance due to the effect of environments was higher than the variances due to genotypes and GEI. The variances ascribed to environments, genotypes, and GEI accounted for 45.79%, 12.96%, and 22.56% of the total variation, respectively. These results indicated that in rain-fed wheat genotypes under study, grain yield was more controlled by the environment than by genetics. AMMI biplot analysis demonstrated that E2 was the main contributor to the GEI given that it was located farthest from the origin. Furthermore, E2 was unstable yet recorded the highest yield. Genotype G47 contributed highly to the GEI sum of squares considering that it was also located far from the origin. Genotypes G12 and G18 were relatively stable because they were situated close to the origin. Their position indicated that they had minimal interaction with the environment. Genotype 47 was the highest-yielding genotype but was unstable, whereas G34 was the lowest-yielding genotype and was unstable.

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Keywords: Genotype–environment interaction, additive main effects and multiplicative interaction analysis, genetics, stability, Triticum aestivum L.

DOI: https://doi.org/10.54910/sabrao2021.53.4.5

Biochemical and cytological features of onion bulbs and leaves collected from various ecogeographical origins

Nemtinov VI, Kostanchuk YN, Pashtetskiy VS, Motyleva SM, Bokhan AI, Caruso G, Katskaya AG, Timasheva LA, Pekhova OA


SUMMARY

Allium cepa L. genotypes with different ecogeographical origins revealed the highest nutritional values when grown in Crimea, Russia. However, their environmental adaptation should be further investigated. This research was performed during 2016–17 to evaluate the qualitative, antioxidant, and mineral composition characteristics of 15 onion genotypes, of which four originated from the Federal State Budget Scientific Institution, Research Institute of Agriculture, Crimea, Russia. Sweet onion genotypes exhibited high ascorbic acid contents of 11.8–27.3 mg/100 g. However, some genotypes had a narrow range of ascorbic acid content (11.8–21.1 mg/100 g) due to their different ecogeographical origins and proved to be appropriate for industrial processing because of their spicy taste. Intensely colored bulbs had a high content of polyphenols (anthocyanins), which are known to have a positive influence on human health. Electronic microscopic assays revealed the morphological characteristics of A. cepa L. genotype leaves and demonstrated the differences in epidermal structure and adaptability potential. The 12 main mineral macro- and microelements with the highest contents in onion leaves were analyzed. The hyperaccumulator genotype with the highest leaf macro- and microelement content could be used to address mineral element deficiencies in humans. In onion genotype leaves, the contents of the mineral elements followed the order of Zn > Fe > Si > Na > Р > Cl > Mo > Mg > S > Ca > Cu > K. The biochemical analysis of 13 onion cultivars showed that the majority exceeded the standard values of dry matter and sugars (mono- and disaccharides) by 13% and 46%, 11% and 48%, and 36% and 150%. In onion genotypes, leaf surface microstructure was specific, and the largest stomata corresponded to the most productive cultivars.

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Date Published: December 2021

Keywords: Onion, bulb (Allium cepa L.), ascorbic acid, leaves, electronic microscopy scanning, energy dispersion X-ray analysis, mineral elements

DOI: https://doi.org/10.54910/sabrao2021.53.4.1

Bio-catharantin effects on phenotypic traits and chromosome number of shallots (Allium Cepa L. var. Ascalonicum ‘Tajuk’)

A.T. BILLA, S.S. LESTARI, B.S. DARYONO, and A.S. SUBIASTUTI

SUMMARY

The seasonal production of Allium cepa var. ascalonicum causes a rise in its demand during the offseason. Consumers mostly prefer onion cultivars like the ‘Super Philip’, because of their high productivity, large and round bulbs, shiny appearance, and less spicy taste. In plant breeding, polyploidy induction through mutagens is a technique often used to produce shallot cultivars of better quality. Bio-Catharantin from the leaf extract of Catharanthus roseus L. is used as a polyploid induction agent instead of colchicine. The latest study aimed to determine the effect of BioCatharantin concentration (0.2% and 0.4%) on phenotypic traits (plant height, bulb mass, and the number of bulbs), and the chromosome number to determine the minimum concentration that could cause polyploidization in shallots. The research was conducted from December 2020 to February 2021 in a greenhouse in Madurejo, Prambanan, and the Laboratory of Genetics and Breeding, Faculty of Biology, Gadjah Mada University, Indonesia. Bio-Catharantin concentration did not affect plant height which was comparable with the control. Both treatments caused an increase in bulb mass up to 37.7 and 41.76 g at the concentrations of 0.2% and 0.4%, respectively, compared with the control (31.47 g). The number of bulbs increased up to 10.6 and 9.8 g for 0.2% and 0.4% concentrations, respectively, compared with 8.8 in the control. The ploidy level of cells was increased from 2n (16) to 3n (24) at 2% and 4n (32) at 4% Bio-Catharantin.

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DOI: http://doi.org/10.54910/sabrao2022.54.2.11

Isolation and diagnosis of cadmium-resistant bacteria and its potential phytoremediation with the broad bean plant

I.A. ABED, A. MARZOOG, A.M.S. ADDAHERI, and M.H. AL-ISSAWI

Results of the study proved that phytoremediation can be a promising technique to treat cadmium (Cd)-contaminated soil. Four bacteria types were isolated from the soil; two are autotrophic and others are heterotrophic. Autotrophic bacteria were dominant in soils with 42 mg Cd Kg-1. The total count and diversity of both bacteria types decreased with the increase of Cd in media and reached their minimum limit of tolerance at 60 mg Cd L-1 in terms of the heterotrophic bacteria, while the minimum limit of tolerance in the case of autotrophic bacteria was at 110 mg Cd L-1. The four isolates can form biofilms that ranged in thickness between 2.8–4.3 mm. The tolerant isolates belong to Rhizobium leguminosarum, Pseudomonas fluorescens, Actinobacteria, and Corynebacterium. Shoot and dry weight significantly varied according to the changes in Cd concentrations and isolate types. The level in either shoot or root exceeded critical levels, however, its concentration was higher in the root compared with the shoot. The effect of Cd on broad bean plants began at 80 and 100 mg Cd L-1. The broad bean plant was resistant to growing in the contaminated area by Cd even at 120 mg Cd Kg1DW. The presence of heterotrophic bacteria was noticeably useful for autotrophic bacteria, as well as, for enhancing Cd resistance. The study showed that cooperative phytoremediation could be a safe and active technique to apply in the field soil contaminated with heavy metals.

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Date published: June 2022

Keywords: Broad bean, cadmium, heavy metals, pollution, resistant bacteria 

DOI: http://doi.org/10.54910/sabrao2022.54.2.17

Gamma-ras and microwave irradiation influence on GUAR (Cyamopsos tetragonoloba: I – markers assisted selection for responding to mutagenic agents

K.A.M. KHALED, F.M. SULTAN, and C.R. AZZAM

SUMMARY

The recent investigation was carried out to determine the effect of different gamma-ray doses and 900 W (2450 MHz) microwave radiation with various exposure times, separately or in combinations, on the yield, yield components, and chemical properties of guar (Cyamopsis tetragonoloba), as well as, to detect variation induction. The cDNA-SCoT technique was used to obtain molecular markers related to some traits. SSR technique was used to sequence the target fragment related to plant height. Gamma-ray doses of 150 and 250 Gy alone, and in combination with 900 W microwaves irradiation applied with different duration or time span (1, 2, 3, and 4 min) influenced the plant height significantly, as well as, number of tillers plant-1 and fresh and dry forage yield, and fresh and dry leaf stem-1ratio. In the second sample, seed yield at harvest time, e.g., pods plant-1, weight of pods plant1 , whole plant dry weight, number of seeds pod-1, length of pod, 100-seed weight, and seeds yield were affected by irradiation with different and varied responses. In the M1 generation, the 18 SCoT primers produced 327 bands ranging between 151–2895 bp in size, out of which 282 were polymorphic (86.24%). In the M2, the 18 SCoT primers produced 328 bands ranging between 2122661 bp in size, out of which 299 were polymorphic (91.16%). The M1 and M2 generations exhibited 89 positive and 39 negative bands, which could be used as marker assisted-selection in response to treated guar plants with different gamma ray doses, separately or in combinations with microwave treatments.

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Date published: June 2022

Keywords: Gamma irradiation, microwave heating, guar grain, yield and yield components, quality analysis, SCoT 

DOI: http://doi.org/10.54910/sabrao2022.54.2.10

Drought effects on mineral composition of the leaves and seeds of Amaranthus tricolor and Amaranthus cruentus

N.V. TETYANNIKOV, S.M. MOTYLEVA, М.S. GINS, N.V. КOZAK, D.V. PANISCHEVA, M.E. MERTVISCHEVA, L.F. KАBASHNIKOVA, I.N. DOMANSKAYA, and Т.S. PILIPOVICH

In global climate change, drought stress is one of the environmental restraining factors that can significantly influence the growth and development of crop plants. Drought stress conditions can also cause undesirable changes in plant physiological and metabolic processes. The influence of soil drought on the mineral composition of leaves and seeds of two species of amaranth (Amaranthus tricolor L. and Amaranthus cruentus L.) with С4-type of photosynthesis was studied through energy dispersive spectrometry (ESD). The recent investigations were carried out during the years 20202022 at the Department of Genofonde and Bioresources of Plants, Federal Scientific Center for Horticulture, Moscow, Russia. The research results showed the leaves of both amaranth with major elements, i.e., K (11.23–15.33), Ca (5.15–7.61), P (3.91–3.92), Mg (2.81–3.36), and Cl (1.86–2.29), whereas, relatively lower values were recorded for Fe (0.05–0.48), and Na (0.07–0.11) mass% respectively. Regarding amaranth plants seed composition, the major elements were K (13.86–13.97), P (7.02–9.76), Mg (3.78–5.64), Ca (3.31–4.78), Cl (2.81–5.30), and Mo (2.80–2.86) mass% respectively. In the species, A. tricolor, a strong correlation was observed between the elements, i.e., S-Cu, Mg-Si, Na-Cu, Na-S, Na-Ca, Na-Si, and Si-S in leaves, while in seeds, these were between CaCu, Mg-Cl, Si-Mn, Ca-Mo, and Cl-Mn. In the other species of amaranth, A. cruentus, the elements viz., Mg-S, Mg-Mo, S-Mo, Mg-Cl, S-Cl, Cl-Mo, Cl-P, P-S, Si-Cl, Ca-Mo, S-Ca, Mg-Ca, Mg-P, P-Mo, and Mg-Si in leaves, while Ni-Cu, Mg-P, Si-P, and Si-Cl in seeds also showed strong relationship. Effects of drought led to a weakening of these ties and the formation of new ones. The accumulation of mineral elements in the leaves of amaranth plants varies from species to species under drought conditions, and A. tricolor cv. Valentina was found most resistant to drought conditions.

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Date published: June 2022

Keywords: Mineral composition, leaves, seeds, drought stress, EDS analysis, Amaranthus tricolor L., Amaranthus cruentus L.

DOI: http://doi.org/10.54910/sabrao2022.54.2.18