Genetic diversity among wild pomegranate (Punica granatum) in Azad Jammu and Kashmir region of Pakistan

BackgroundPomegranate (Punica granatum L.), one of the most important tropical fruits in Azad Jammu and Kashmir regions of Pakistan, is highly valued for its nutrition and medicinal purposes. Although pomegranate is native to this region, the genetic diversity among wild pomegranate accessions is currently unknown. Such information would be vital for germplasm conservation and breeding efforts. In the current study, genetic diversity among forty-eight wild pomegranate accessions collected from different agro-ecological zones of Azad Jammu and Kashmir was assessed using 41 simple sequence repeat (SSR) markers.ResultsThe markers revealed 303 alleles averaging 7.39 alleles per marker. Polymorphic information content ranged from 0.12 (PGCT093B) to 0.88 (Pom006), with a mean of 0.54. The average genetic distance (GD) across all genotypes was 0.52, and was lowest between Chattar Class and Thorar genotypes (GD = 0.27), but highest between Khun Bandway and Akhor Ban (GD = 0.74). A neighbor-joining dendrogram separated the genotypes into three major clusters, with further sub-clustering within each cluster.ConclusionsOverall, the results presented here show significant genetic diversity among wild pomegranate accessions in Azad Jammu and Kashmir region of Pakistan. These accessions present a valuable genetic resource to breeding and cultivar improvement programs within the region.How to cite: Aziz S, Firdous S, Rahman H, et al. Genetic diversity among wild pomegranate (Punica granatum) in Azad Jammu and Kashmir region of Pakistan. Electron J Biotechnol 2020;46. https://doi.org/10.1016/j.ejbt.2020.06.002.     

Development and application of two novel functional molecular markers of BADH2 in rice

BackgroundFragrance is one of the most important quality traits in rice, and the phenotype is attributed to the loss-of-function betaine aldehyde dehydrogenase (BADH2) gene. At least 12 allelic variations of BADH2 have been identified, and some of these have been applied to rice fragrance breeding using traditional molecular markers and Sanger sequencing techniques. However, these traditional methods have several limitations, such as being very expensive, imprecise, inefficient, and having security issues. Thus, a new molecular marker technology must be developed to improve rice fragrance breeding.ResultsIn this study, more than 95% of the cultivated fragrant rice varieties belonged to a 7-bp deletion in exon 2 (badh2-E2) or an 8-bp deletion and 3-bp variation in exon 7 (badh2-E7). Both allelic variations resulted in the loss of function of the badh2 gene. We developed two novel SNP molecular markers, SNP_badh2-E2 and SNP_badh2-E7, related to the alleles. Their genotype and phenotype were highly cosegregated in the natural variation of rice accessions, with 160 of the 164 fragrant rice varieties detected with the two markers. These markers cosegregated with the fragrance phenotype in the F₂ population.ConclusionsTwo functional SNP molecular markers of badh2-E2 and badh2-E7 allelic variations were developed. These functional SNP molecular markers can be used for genotype and genetic improvement of rice fragrance through marker-assisted selection and will significantly improve the efficiency of fragrant rice breeding and promote commercial molecular breeding of rice in the future.How to cite: Li W, Zeng X, Li S, et al. Development and application of two novel functional molecular markers of BADH2 in rice. Electron J Biotechnol 2020;46. https://doi.org/10.1016/j.ejbt.2020.04.004.     

Genome-wide development of polymorphic microsatellite markers and their application in peanut breeding program

BackgroundCultivated peanut (Arachis hypogaea. L) represents one of the most important oil crops in the world. Although much effort has been expended to characterize microsatellites or Simple Sequence Repeats (SSRs) in peanut, the quantity and quality of the markers in breeding applications remain limited. Here, genome-wide SSR characterization and marker development were performed using the recently assembled genome of the cultivar Tifrunner.ResultsIn total, 512,900 microsatellites were identified from 2556.9-Mb genomic sequences. Based on the flanking sequences of the identified microsatellites, 7757 primer pairs (markers) were designed, and further evaluated in the assembled genomic sequences of the tetraploid Arachis cultivars, Tifrunner and Shitouqi, and the diploid ancestral species, A. duranensis and A. ipaensis. In silico PCR analysis showed that the SSR markers had high amplification efficiency and polymorphism in four Arachis genotypes. Notably, nearly 60% of these markers were single-locus SSRs in tetraploid Arachis species, indicating they are more specific in distinguishing the alleles of the A and B sub-genomes of peanut. In addition, two markers closely related with purple testa color and 27 markers near to FAD2 genes were identified, which could be used for breeding varieties with purple testa and high-oleic acid content, respectively. Moreover, the potential application of these SSR markers in tracking introgressions from Arachis wild relatives was discussed.ConclusionsThis study reported the development of genomic SSRs from assembled genomic sequences of the tetraploid Arachis Tifrunner, which will be useful for diversity analysis, genetic mapping and functional genomics studies in peanut.How to cite: Ma J, Zhao Y, Chen H, et al. Genome-wide development of polymorphic microsatellite markers and their application in peanut breeding program. Electron J Biotechnol 2020;44. https://doi.org/10.1016/j.ejbt.2020.01.004.     

Identification of genetic variants the CCKAR gene and based on body measurement and carcass quality characteristics in Qinchuan beef cattle (Bos taurus)

BackgroundThis study aimed to explore genetic polymorphisms of the CCKAR gene and their relationship with the growth and development of Qinchuan cattle which could be used as molecular markers for the improvement of the breeding of Qinchuan cattle.ResultsHere, we have identified seven single nucleotide polymorphisms (SNPs) at loci g. 1463 C>G; g. 1532 T>A; g. 1570 G>A; g. 1594 C>A; g. 1640 T>C; g. 1677 G>C; and g. 1735 C>T in the coding region of the bovine CCKAR gene. The frequencies identified on allelic and genotypic characteristics have shown that all seven SNPs diverged from the Hardy-Weinberg-Equilibrium. The SNP2, SNP3, SNP6 and SNP7 had the lowest polymorphism information content values, and remaining SNPs were found to be moderate (0.25 < PIC < 0.50). The genotype CG in SNP1 at loci g.1463 C>G had the greatest association with WH, HW, CD and CCF, while the genotype TA at the very same loci was associated with BFT, ULA and IMF content in Qinchuan cattle. The CCKAR gene expression level in adipose tissue, small intestine, liver and skeleton muscle was found to be higher, whereas, the expression level of mRNA in organs of other digestive system including reticulum, abomasum and omasum was moderate. Some expression of CCKAR mRNA was found in the large intestine, kidney and rumen.ConclusionsIn summary, our finding suggested that the CCKAR gene could be used as a potential candidate for the improvement of carcass quality and body measurements of Qinchuan cattle.How to citeNurgulsim K, Raza SHA, Khan R, et al. Identification of genetic variants the CCKAR gene and based on body measurement and carcass quality characteristics in Qinchuan beef cattle (Bos taurus). Electron J Biotechnol 2021;51. https://doi.org/10.1016/j.ejbt.2021.02.001     

Bovine TMEM95 gene: Polymorphisms detecting in five Chinese indigenous cattle breeds and their association with growth traits

BackgroundTransmembrane protein 95 (TMEM95) plays a role in male fertility. Previous studies showed that genes with a significant impact on reproductive traits can also affect the growth traits of livestock. Thus, we speculated that the genetic variation of TMEM95 gene may have effects on growth traits of cattle.ResultsTwo SNPs were genotyped. The rs136174626 and rs41904693 were in the intron 4 and 3′-untranslated region, respectively. The linkage disequilibrium analysis illustrated that these two loci were not linked. The rs136174626 was associated with six growth traits of Nanyang cattle, four traits of Luxi cattle, and three traits of Ji’an cattle. For rs41904693 locus, the GG individuals had greater body height and abdominal girth in Ji’ an cattle than TT and TG individuals. In Jinnan cattle, GG and TT individuals had greater body height, height at hip cross, body length, and heart girth than TG individuals. The potential splice site prediction results suggest that the rs136174626 may influence the splicing efficiency of TMEM95, and the miRNA binding site prediction results showed that the rs41904693 may influence the expression of TMEM95 by affecting the binding efficiency of Bta-miR-1584 and TMEM95 3′-UTR.ConclusionsThe findings of the study suggested that the two SNPs in TMEM95 could be a reliable basis for molecular breeding in cattle.How to cite: Guo X, Zhang S, Yang H, et al. Bovine TMEM95 gene: Polymorphisms detecting in five Chinese indigenous cattle breeds and their association with growth traits. Electron J Biotechnol 2021;51. https://doi.org/10.1016/j.ejbt.2021.03.004     

Production of wheat-doubled haploids resistant to eyespot supported by marker-assisted selection

BackgroundWheat is one of the most important crops cultivated all over the world. New high-yielding cultivars that are more resistant to fungal diseases have been permanently developed. The present study aimed at the possibility of accelerating the process of breeding new cultivars, resistant to eyespot, by using doubled haploids (DH) system supported by marker-assisted selection.ResultsTwo highly resistant breeding lines (KBP 0916 and KBH 4942/05) carrying Pch1 gene were crossed with the elite wheat genotypes. Hybrid plants of early generations were analyzed using endopeptidase EpD1 and two SSR markers linked to the Pch1 locus. Selected homozygous and heterozygous genotypes for the Pch1-linked EpD1b allele were used to produce haploid plants. Molecular analyses were performed on haploids to identify plants possessing Pch1 gene. Chromosome doubling was performed only on haploid plants with Pch1 gene. Finally, 65 DH lines carrying eyespot resistance gene Pch1 and 30 lines without this gene were chosen for the eyespot resistance phenotyping in a field experiment.ConclusionsResults of the experiment confirmed higher resistance to eyespot of the genotypes with Pch1 in comparison to those without this gene. This indicates the efficiency of selection at the haploid level.How to cite: Wiśniewska H, Majka M, Kwiatek M, et al. Production of wheat doubled haploids resistant to eyespot supported by marker-assisted selection. Electron J Biotechnol 2019;37. https://doi.org/10.1016/j.ejbt.2018.10.003     

Genetic diversity and differentiation of nine populations of the hard clam (Meretrix petechialis) assessed by EST-derived microsatellites

BackgroundMeretrix petechialis is one of the commercially important marine bivalves. In this study, we selected six highly polymorphic EST-derived microsatellite markers to assess the genetic diversity and population differentiation on nine wild populations of Meretrix petechialis.ResultsThe number of alleles detected per locus ranged from 4 to 30 (mean N_A = 27.5) with a total of 165 alleles. The mean value of observed and expected heterozygosities varied from 0.717 to 0.861 and from 0.797 to 0.856, respectively. Meanwhile, the result of Neighbor-joining and overall F_ST = 0.214 (P < 0.01) reveled that M. petechialis populations from GX are the farthest populations, a certain degree of genetic variation among individuals in each population and the genetic differentiation is significant.ConclusionsGX population has high genetic diversity among individual, and there are certain differences in genetic characteristics among different populations. This study will provide a basis for the domestication and cultivation of genetic diversity of M. petechialis population and the protection of clam germplasm resources.How to citeXu Q, Zheng J, Yan X, et al. Genetic diversity and differentiation of nine populations of the hard clam (Meretrix petechialis) assessed by EST-derived microsatellites. Electron J Biotechnol 2020;48. https://doi.org/10.1016/j.ejbt.2020.09.003     

Analysis of the population structure and genetic diversity of the red swamp crayfish (Procambarus clarkii) in China using SSR markers

BackgroundProcambarus clarkii produces high-quality, delicious meat that is high in protein, low in fat, and rich in calcium and phosphorus. It has become an important aquatic resource in China. Our objectives are (i) to analyze the level of genetic diversity of P. clarkii populations; (ii) to explore the genetic differentiation (Gst); and (iii) to propose appropriate strategies for the conservation.ResultsIn this study, Shannon's index (I) and Nei's gene diversity index (H) for P. clarkii were high (I = 0.3462 and H = 0.2325 on average and I = 0.6264, H = 0.4377 at the species level) based on the SSR markers. The expected heterozygosity value of 17 microsatellite loci in 25 crayfish populations was 0.9317, the observed heterozygosity value was 0.9121, and the observed number of alleles per locus was 2.000; and the effective number of alleles per locus was 1.8075. Among the P. clarkii populations, the inbreeding coefficient within populations (Fis) was 0.2315, overall inbreeding coefficient (Fit) was 0.4438, genetic differentiation coefficient among populations (Fst) was 0.3145 and gene differentiation (Gst) was 0.4785 based on SSR analyses. The cluster analysis results obtained by unweighted pair-group method with arithmetic mean (UPGMA) analysis, principal coordinate analysis (PCoA) and STRUCTURE analysis were similar. A mantel test showed that the isolation-by-distance pattern was not significant.ConclusionsThe high Gst among P. clarkii populations is attributed to genetic drift and geographic isolation. The results indicated that more P. clarkii populations should be collected when formulating conservation and aquaculture strategies.How to citeLiu F, Qu Y-K, Geng C, et al. Analysis of the population structure and genetic diversity of the red swamp crayfish (Procambarus clarkii) in China using SSR markers. Electron J Biotechnol 2020;47. https://doi.org/10.1016/j.ejbt.2020.06.007.     

Bioleaching of realgar nanoparticles using the extremophilic bacterium Acidithiobacillus ferrooxidans DLC-5

BackgroundThis paper presents micro- and nano-fabrication techniques for leachable realgar using the extremophilic bacterium Acidithiobacillus ferrooxidans (A. ferrooxidans) DLC-5.ResultsRealgar nanoparticles of size ranging from 120 nm to 200 nm were successfully prepared using the high-energy ball mill instrument. A. ferrooxidans DLC-5 was then used to bioleach the particles. The arsenic concentration in the bioleaching system was found to be increased significantly when compared with that in the sterile control. Furthermore, in the comparison with the bioleaching of raw realgar, nanoparticles could achieve the same effect with only one fifth of the consumption.ConclusionEmphasis was placed on improving the dissolvability of arsenic because of the great potential of leachable realgar drug delivery in both laboratory and industrial settings.How to cite: Xu R, Song P, Wang J, et al. Bioleaching of realgar nanoparticles using the extremophilic bacterium Acidithiobacillus ferrooxidans DLC. Electron J Biotechnol 2019;38. https://doi.org/10.1016/j.ejbt.2019.01.001.     

Polymorphism of bovine lipocalin-2 gene and its impact on milk production traits and mastitis in Holstein Friesian cattle

BackgroundMastitis is one of the most serious diseases of dairy cattle, causing substantial financial losses. While predisposition to reduced somatic cell count in milk has been considered for in cattle breeding programs as the key indicator of udder health status, scientists are seeking genetic markers of innate immune response, which could be helpful in selecting cows with improved immunity to mastitis. Lipocalin-2 (LCN2) is a protein involved in the response of the immune system by eliminating iron ions which are necessary for the growth of pathogenic bacteria, so LCN2 may be considered as a natural bacteriostatic agent and could become a marker of infection.ResultsA total of five SNPs were identified in LCN2 gene (one in the promoter, three in exon 1, and one in intron 1). A single haplotype block was identified. The locus g.98793763G > C was found to have a significant impact on protein levels in milk, and alleles of this locus were identified to have a significant positive dominance effect on this trait. None of the four analysed loci had a statistically significant impact on the milk yield, fat levels in milk or the somatic cell score. LCN-2 gene had no significant impact on the incidence of mastitis in the cows.ConclusionsAlthough the identified SNPs were not found to have any impact on the somatic cell count or the incidence of mastitis in cows, it seems that further research is necessary, covering a larger population of cattle, to confirm the association between lipocalin-2 and milk production traits and mastitis.How to cite: Pokorska J, Piestrzyńska-Kajtoch A, Kułaj D, et al. Polymorphism of bovine lipocalin-2 gene and its impact on milk production traits and mastitis in Holstein Friesian cattle. Electron J Biotechnol 2019;40. https://doi.org/10.1016/j.ejbt.2019.04.004     

Effective diffusion coefficients and bioconversion rates of inhibitory compounds in flocs of Saccharomyces cerevisiae

BackgroundFermentation strategies for bioethanol production that use flocculating Saccharomyces cerevisiae yeast need to account for the mechanism by which inhibitory compounds, generated in the hydrolysis of lignocellulosic materials, are tolerated and detoxified by a yeast floc.ResultsDiffusion coefficients and first-order kinetic bioconversion rate coefficients were measured for three fermentation inhibitory compounds (furfural, hydroxymethylfurfural, and vanillin) in self-aggregated flocs of S. cerevisiae NRRL Y-265. Thièle-type moduli and internal effectiveness factors were obtained by simulating a simple steady-state spherical floc model.ConclusionsThe obtained values for the Thiéle moduli and internal effectiveness factors showed that the bioconversion rate of the inhibitory compounds is the dominant phenomenon over mass transfer inside the flocs.How to cite: Landaeta R, Acevedo F, Aroca G. Effective diffusion coefficients and bioconversion rates of inhibitory compounds in flocs of Saccharomyces cerevisiae. Electron J Biotechnol 2019;42. https://doi.org/10.1016/j.rjbt.2019.08.001     

Enhanced pyruvic acid yield in an osmotic stress-resistant mutant of Yarrowia lipolytica

BackgroundPyruvic acid (PA), a vital α-oxocarboxylic acid, plays an important role in energy and carbon metabolism. The oleaginous yeast Yarrowia lipolytica (Y. lipolytica) has considerable potential for the production of PA. An increased NaCl concentration reportedly increases the biomass and PA yield of Y. lipolytica.ResultsTo increase the yield of PA, the NaCl-tolerant Y. lipolytica A4 mutant was produced using the atmospheric and room temperature plasma method of mutation. The A4 mutant showed growth on medium containing 160 g/L NaCl. The PA yield of the A4 mutant reached 97.2 g/L at 120 h (0.795 g/g glycerol) in a 20-L fermenter with glycerol as the sole carbon source, which was 28.9% higher than that of the parental strain.ConclusionThe PA yield from Y. lipolytica can be improved by increasing its NaCl tolerance.How to cite: Yuan W, Lin X, Zhong S, et al. Enhanced pyruvic acid yield in an osmotic stress-resistant mutant of Yarrowia lipolytica. Electron J Biotechnol 2020;44. https://doi.org/10.1016/j.ejbt.2020.01.002.     

Development and application of KASP marker for high throughput detection of AhFAD2 mutation in peanut

BackgroundCultivated peanut (Arachis hypogaea L.) is a major oilseed crop worldwide. Fatty acid composition of peanut oil may affect the flavor and shelf life of the resulting food products. Oleic acid and linoleic acid are the major fatty acids of peanut oil. The conversion from oleic acid to linoleic acid is controlled by the Δ12 fatty acid desaturase (FAD) encoded by AhFAD2A and AhFAD2B, two homoeologous genes from A and B subgenomes, respectively. One nucleotide substitution (G:C → A:T) of AhFAD2A and an “A” insertion of AhFAD2B resulted in high-oleic acid phenotype. Detection of AhFAD2 mutation had been achieved by cleaved amplified polymorphic sequence (CAPS), real-time polymerase chain reaction (qRT-PCR) and allele-specific PCR (AS-PCR). However, a low cost, high throughput and high specific method is still required to detect AhFAD2 genotype of large number of seeds. Kompetitive allele specific PCR (KASP) can detect both alleles in a single reaction. The aim of this work is to develop KASP for detection AhFAD2 genotype of large number of breeding materials.ResultsHere, we developed a KASP method to detect the genotypes of progenies between high oleic acid peanut and common peanut. Validation was carried out by CAPS analysis. The results from KASP assay and CAPS analysis were consistent. The genotype of 18 out of 179 BC₄F₂ seeds was aabb.ConclusionsDue to high accuracy, time saving, high throughput feature and low cost, KASP is more suitable for determining AhFAD2 genotype than other methods.     

Effects of exogenous γ-Aminobutyric acid on the regulation of respiration and protein expression in germinating seeds of mungbean (Vigna radiata) under salt conditions

Backgroundγ-Aminobutyric acid (GABA) bypasses the TCA cycle via GABA shunt, suggesting a relationship with respiration. However, little is known about its role in seed germination under salt conditions.ResultsIn this study, exogenous GABA was shown to have almost no influence on mungbean seed germination, except 0.1 mM at 10 h, while it completely alleviated the inhibition of germination by salt treatment. Seed respiration was significantly inhibited by 0.1 and 0.5 mM GABA, but was evidently enhanced under salt treatment, whereas both were promoted by 1 mM GABA alone or with salt treatment. Mitochondrial respiration also showed a similar trend at 0.1 mM GABA. Moreover, proteomic analysis further showed that 43 annotated proteins were affected by exogenous GABA, even 0.1 mM under salt treatment, including complexes of the mitochondrial respiratory chain.ConclusionsOur study provides new evidence that GABA may act as a signal molecule in regulating respiration of mungbean seed germination in response to salt stress.How to citeJi J, Shi S, Chen W, et al. Effects of exogenous γ-Aminobutyric acid on the regulation of respiration and protein expression in germinating seeds of mungbean (Vigna radiata) under salt conditions. Electron J Biotechnol 2020;47. https://doi.org/10.1016/j.ejbt.2020.05.005     

Study of capability of nanostructured zero-valent iron and graphene oxide for bioremoval of trinitrophenol from wastewater in a bubble column bioreactor

BackgroundBioremoval of phenolic compounds using fungi and bacteria has been studied extensively; nevertheless, trinitrophenol bioremediation using modified Oscillatoria cyanobacteria has been barely studied in the literature.ResultsAmong the effective parameters of bioremediation, algal concentration (3.18 g·L⁻¹), trinitrophenol concentration (1301 mg·L⁻¹), and reaction time (3.75 d) were screened by statistical analysis. Oscillatoria cyanobacteria were modified by starch/nZVI and starch/graphene oxide in a bubble column bioreactor, and their bioremoval efficiency was investigated. Modifiers, namely, starch/zero-valent iron and starch/GO, increased trinitrophenol bioremoval efficiency by more than 10% and 12%, respectively, as compared to the use of Oscillatoria cyanobacteria alone.ConclusionsIt was found that starch/nano zero-valent iron and starch/GO could be applied to improve the removal rate of phenolic compounds from the aqueous solution.How to cite: Bavandi R, Emtyazjoo M, Saravi HN, et al. Study of nano-structure zero-valent iron and graphene-oxid capability onbioremoval of trinitrophenol from wastewater in a bubble column bioreactor. Electron J Biotechnol 2019;39. https://doi.org/10.1016/j.ejbt.2019.02.003.