15 research outputs found
Cloning Abiotic Stress Associated Genes and Agrobacterium Tumefaciens Mediated Transformation of Selected Tropical Maize
No full textDrought and salinity are the main abiotic constraints of maize production. The
genes confer tolerance to abiotic stresses can be isolated, cloned and introduced
into important crops such as maize. The Annexin1 (AnnAt1) and Annexinp35
have been highly associated with water stress tolerance. AnnAt1 is induced by
various stresses including water and oxidative stress and has been found to
have peroxidase activity. Annexin p35 is a maize gene that also has peroxidase
activity and plays a role in exocytosis, calcium transport and regulation. The
Na+/H+ antiporter (NHX1) gene is involved in compartmentalization of Na+
into the vacuoles and therefore playing an important role in salt tolerance.
However these genes have not been reported in transformation of tropical
maize and its response to water stress. Annexinp35 has never been cloned and
used in transformation of any crop. The objectives of this study was to isolate
and clone the maize Annexinp35 gene and transform tropical maize with the
Annexinp35, AnnAt1 and NHX1 genes conferring drought and salt tolerance
via Agrobacterium tumefaciens and screening for stable integration and
expression of the transgenes in T0 transgenic plants. An initial assessment of
maize response to callus induction was performed using mature embryos, shoot
tips and leaf segments and different levels of 2,4-Dichlorophenoxyacetic acid
(2,4-D). AnnexinP35 gene was isolated from egg cells of maize and cloned
into pNOV2819 vector, the AnnAt1 gene in pROK2 vector was sub-cloned into
pNOV2819 vectors with it is promoter and terminator, the PMI gene from
pNOV2819 vector was sub-cloned in to pCAMBIA-NHX1 vector which
carries the salt tolerant gene NHX1 gene. Drought and salt tolerant genes were
engineered into Sudanese maize genotypes, These genotypes are both striger
resistant and most preferred by farmers in Sudan. The drought and salt
tolerance genes were introduced to Sudanese maize using Agrobacterium
tumefaciens method. The experiment was carried out using random complete
block design. Transformation frequency and efficiency were assessed by using
mannose as selectable agent. Transformation frequency and efficiency were
found to be genotypic dependant. Transformation frequency and efficiency
were evaluated for all the genotypes used in the study. Shoot tips gave the
highest callus induction frequency among all the genotypes used while mature
embryos gave the lowest callus induction frequency. The highest
transformation frequency for the gene construct pCAMBIA-NHX1/PMI was
observed in STR136 (8.03%). IL15 and Mojatamma-45 had TF of 2.05% and
1.75% respectively while the highest TE was observed in IL15 (1.13%). The
highest transformation frequency for the vector pNOV2819-
ASARZMANNp35 was observed in Hudiba-2 (31.78%) while the lowest TF
was observed in Mojtamma-45 (5.38%). The highest TE was observed for
Giza-2 (1.00%). The gene construct pNOV2819-AnnAt1 gave highest
transformation frequency in the Inbred local-5 (23.98%) and lowest TF in IL1
(3.75%). The highest TE was observed Giza-2 (2.48%) for the same gene
construct. Drought tolerant lines generated will be available to the maize
breeders to transfer the trait to lines that have high yield but lack this trait
Agrobacterium Tumefaciens-Mediated Transformation of Sudanese Maize Genotypes Using NPK1 Gene for Enhancing Drought Stress Tolerance
No full textDrought is one of the most important abiotic factor affecting maize production
worldwide. Agrobacterium-mediated gene transfer technique has been established as a
versatile way of improving important crops for tolerance to biotic and abiotic factors.
Through this technique, the drought tolerance gene, NPK1, has been used in the
transformation of temperate maize after its isolation and characterization from
tobacco. Recovered transgenic events were observed to have enhanced tolerance to
water stress. The accelerated adoption of the transformation technique in Africa, and
indeed in Sudan, will depend on the ease with which transgenes of agronomic
importance can be integrated into appropriate germplasms. This study aimed at
screening important Sudanese maize inbred lines and open pollinated varieties
(OPVs) for transformability via the integration of the NPK1 gene. Eight inbred lines
and three OPVs were evaluated. A188 was used as the standard inbred line check
while KAT was used as the local OPV check. Freshly isolated immature embryos of
maize were inoculated with Agrobacterium strain EHA 101 harbouring the plasmid
pSHX004 in LS infection media for 5 minutes and then co-cultivated on LS cocultivation
media for 3 days. Embryos were then transferred to selection media
supplemented with 250mg/l cefotaxime and 1.5mg/L bialaphos. After two weeks on
this media, calli were subcultured on selection media containing 3.0 mg/L bialaphos
for 4 weeks. Bialaphos resistant callus events were then transferred to maturation
media supplemented 3mg/L bialaphos for 2 weeks before transferring to shooting
media. Shoots were then transferred to rooting media. Plantlets with well-formed root
system were transferred from the in vitro environment to green house for hardening.
Hardened plantlets were transplanted to soil in the greenhouse and maintained till they
set seeds. To confirm the presence of the transgene, PCR analysis was done on
putative transgenic plants using the Bar primers. Out of a total of 4401 immature
embryos from the 13 genotypes infected, 327 survived selection in bialaphos.
Bialaphos resistant calli emerged 3-4 weeks after selection. IL3, IL15, Hudiba-2, IL1,
IL38, Hudiba-1, A188 and KAT produced compact calli from their scutella surfaces
while IL28, IL42, IL43, Mojtamaa-45 and IL16 established watery nonembryogenic
calli. Statistically significant differences (p<0.05) were observed between the
genotypes with respect to transformation frequency (TF). IL3 was identified as the
most amenable to transformation with a TF of 31.7% and proved to be superior to
A188, which recorded a TF of 5.82%. Hudiba-2 was identified as the most
transformable OPV with a TF of 8.7% compared to that of 7.3% for KAT. IL1 and
Mojatamaa-45 proved to be poor responders to transformation with TFs of 2.5% and
1.7%, respectively. Putative transgenics were recovered from IL3, IL15, Hudiba-2,
IL1, IL38, Mojatamaa-45, A188 and KAT. The frequency of regeneration of
bialaphos resistant shoots varied from 6.9% for IL38 to 100% for Mojtamaa-45. PCR
analysis indicated a 540bp fragment in the DNA extracts from transgenic R₁ plants.
Transformation efficiency (TE) was found to depend on the genotype used. The
highest TE was observed for IL3 (3.7%), while the lowest TE of 0.0% was observed
in IL42 IL43, IL16 and IL28. Various abnormalities were observed in putative
transformants including dwarfism, tussel seed and lack of ear. However, plants grew
to maturity and were able to establish seeds in spite of these abnormalities. In
conclusion, the inbred line IL3 and the OPV Hudiba-2 proved to be the most
amenable Sudanese genotypes to A. tumefaciens-mediated transformation. Future
research in maize improvement through biotechnologies such as tissue culture and
genetic transformation should be focused on these good responders
Callus Induction and Plant Regeneration from Immature Embryos of Sweet Sorghum (Sorghum bicolor Moench)
No full textThis study sought to establish the effect of genotype and auxin concentration on callus induction from different sweet sorghum genotypesBackground and Objective: Regeneration of plant through tissue culture technique is a critical process in transformation of plants. This study sought to establish the effect of genotype and auxin concentration on callus induction from different sweet sorghum genotypes (IESV92008DL, IESV92001DL, IESV92021DL, ICSV700 and ICSV93048). Materials and Methods: In this study, MS medium supplemented with five levels of the hormone 2,4-D (0, 1, 2, 4 and 6 mg LG1) to assess the effect of the hormone 2,4-D on callus induction and regeneration was used. Results: The highest callus induction frequency was observed at 2 mg LG1 2,4-D for all the genotypes, IESV92008 gave the highest callus induction frequency among all the genotypes at 2 mg LG1 2,4-D. The lowest callus induction frequency was observed 0 and 6 mg LG1 for all the genotypes, the two genotypes ICSV700 and IESV92021 were observed to give the lowest callus induction frequency among all the genotypes used in this study. Embryogenic callus induction frequency was observed to be higher at 2 mg LG1 2,4-D and the genotype IESV92008 gave the highest callus induction frequency among all the genotypes used. Induction of shoots was achieved in MS medium supplemented with 3 mg LG1 6BA and 1 mg LG1 IAA, the highest regeneration efficiency was obtained from the three genotypes ICSV93048, IESV92008 and IESV92001, respectively. Conclusion: This study discovers the importance of the auxin 2,4-D on callus induction and regeneration of sweet sorghum and this will help the researcher to develop protocols for transformation of sweet sorghum
Callus Induction and Plant Regeneration from Immature Embryos of Sweet Sorghum (Sorghum bicolor Moench)
No full textCallus Induction and Plant Regeneration from Immature Embryos of Sweet Sorghum (Sorghum bicolor Moench)
No full text
Potential causal association between gut microbiome and posttraumatic stress disorder
No full textFunding Information: We thank the participants and working staff including the Psychiatric Genomics Consortium Posttraumatic Stress Disorder Working Group, the FinnGen consortium, and the MiBioGen consortium. Publisher Copyright: © 2024, The Author(s).Background: The causal effects of gut microbiome and the development of posttraumatic stress disorder (PTSD) are still unknown. This study aimed to clarify their potential causal association using mendelian randomization (MR). Methods: The summary-level statistics for gut microbiome were retrieved from a genome-wide association study (GWAS) of the MiBioGen consortium. As to PTSD, the Freeze 2 datasets were originated from the Psychiatric Genomics Consortium Posttraumatic Stress Disorder Working Group (PGC-PTSD), and the replicated datasets were obtained from FinnGen consortium. Single nucleotide polymorphisms meeting MR assumptions were selected as instrumental variables. The inverse variance weighting (IVW) method was employed as the main approach, supplemented by sensitivity analyses to evaluate potential pleiotropy and heterogeneity and ensure the robustness of the MR results. We also performed reverse MR analyses to explore PTSD’s causal effects on the relative abundances of specific features of the gut microbiome. Results: In Freeze 2 datasets from PGC-PTSD, eight bacterial traits revealed a potential causal association between gut microbiome and PTSD (IVW, all P < 0.05). In addition, Genus.Dorea and genus.Sellimonas were replicated in FinnGen datasets, in which eight bacterial traits revealed a potential causal association between gut microbiome and the occurrence of PTSD. The heterogeneity and pleiotropy analyses further supported the robustness of the IVW findings, providing additional evidence for their reliability. Conclusion: Our study provides the potential causal impact of gut microbiomes on the development of PTSD, shedding new light on the understanding of the dysfunctional gut-brain axis in this disorder. Our findings present novel evidence and call for investigations to confirm the association between their links, as well as to illuminate the underlying mechanisms.publishersversionpublishe
SARS-CoV-2 vaccination modelling for safe surgery to save lives: data from an international prospective cohort study
No full textBackground: Preoperative SARS-CoV-2 vaccination could support safer elective surgery. Vaccine numbers are limited so this study aimed to inform their prioritization by modelling.
Methods: The primary outcome was the number needed to vaccinate (NNV) to prevent one COVID-19-related death in 1 year. NNVs were based on postoperative SARS-CoV-2 rates and mortality in an international cohort study (surgical patients), and community SARS-CoV-2 incidence and case fatality data (general population). NNV estimates were stratified by age (18-49, 50-69, 70 or more years) and type of surgery. Best- and worst-case scenarios were used to describe uncertainty.
Results: NNVs were more favourable in surgical patients than the general population. The most favourable NNVs were in patients aged 70 years or more needing cancer surgery (351; best case 196, worst case 816) or non-cancer surgery (733; best case 407, worst case 1664). Both exceeded the NNV in the general population (1840; best case 1196, worst case 3066). NNVs for surgical patients remained favourable at a range of SARS-CoV-2 incidence rates in sensitivity analysis modelling. Globally, prioritizing preoperative vaccination of patients needing elective surgery ahead of the general population could prevent an additional 58 687 (best case 115 007, worst case 20 177) COVID-19-related deaths in 1 year.
Conclusion: As global roll out of SARS-CoV-2 vaccination proceeds, patients needing elective surgery should be prioritized ahead of the general population
Global warming and malaria: a call for accuracy
No full textFor more than a decade, malaria has held a prominent place in speculations on the impacts of global climate change. Mathematical models that “predict? increases in the geographic distribution of malaria vectors and the prevalence of the disease have received wide publicity. Efforts to put the issue into perspective1, 2, 3, 4 and 5 are rarely quoted and have had little influence on the political debate. The model proposed by Frank C Tanser and colleagues6 in The Lancet and the accompanying Commentary by Simon Hales and Alistair Woodward7 are typically misleading examples.The relation between climate and malaria transmission is complex and varies according to location,2 yet Tanser et al base their projections on thresholds derived from a mere 15 African locations. Slight adjustments of values assigned to such thresholds and rules can influence spatial predictions strongly.8 The authors invest considerable effort in assessing the sensitivity of their model to climate change scenarios but do not report the internal sensitivities to thresholds and rules. The predictive skill of their model is low (63% sensitivity, 95% CI 61–65%) but they consider projections acceptable if prevalence is projected “to within a month? (presumably +/- 1 month?), thereby biasing their model towards success. A model covering an entire year in a parasite-positive site would always be correct, although in such areas it would be relatively insensitive to climate. By contrast, sites in which transmission is seasonal would provide a more reliable test of accuracy, but estimation is more difficult because climate sensitivity is greater. Furthermore, because parasite clearance in communities is not instantaneous,9 spot samples of parasitaemia on survey dates are not a suitable indicator of the duration of the transmission season. Lastly, “person/months? are unsuitable as a measure of transmission: an extension of season from 1 to 4 months will have more impact than from 10 to 12 months. According to their model, an extension of transmission from 11 to 12 months results in 106 more person/months in a population of 106 people, whereas an extension from 1 to 5 months gives the same increase in a population of 250·000.What Tanser and colleagues have modelled is merely the duration of the transmission season, which they interpret as “heightened transmission? and increased incidence. A greater failing is their reliance on “parasite-ratio studies?. The relations between transmission season and parasite prevalence, and parasite prevalence and clinical disease, are unclear but unlikely to be linear. Moreover, they use 1995 data for human populations, although these are projected to double by 2030. In addition, the proportion living in urban areas—with a specific climate10 and orders of magnitude less malaria transmission11 and 12—is projected to rise from 37% to 53%.13 For all these reasons, we do not accept the model as a “baseline against which interventions can be planned?.It is regrettable that many involved in this debate ignore the rich heritage of literature on the subject. For example, in 1937, in his classic textbook,14 L W Hackett stated: “Everything about malaria is so moulded and altered by local conditions that it becomes a thousand different diseases and epidemiological puzzles. Like chess, it is played with a few pieces, but is capable of an infinite variety of situations?. A pressing question in Hackett's time was the changing distribution of the disease in Europe. On the role of climate, he wrote: “Certainly, climate lays down the broad lines of malaria distribution…Nevertheless, although this is a very simple and plausible explanation…even the early malariologists felt that there was something unsatisfactory about it…malaria has not so much receded as it has contracted, oftentimes toward the north…Thus in Germany it is the northern coast which is still malarious, the south is free…There is, therefore, no climatic reason why (malaria) should have abandoned south Germany or the French Riviera?.We quote Hackett because we feel that the classic components of science—unbiased observation and systematic experimentation—cannot be sidestepped with models that omit many of his chess pieces. Yet Hales and Woodward7 begin by stating: “The present geographical distribution of malaria is explained by a combination of environmental factors (especially climate) and social factors (such as disease-control measures)?. In our opinion, “even the early malariologists? would surely disagree: much of the decline of malaria in Europe took place without control measures during a period when the climate was warming.The text by Hales and Woodward that follows displays a lack of knowledge. Thus, “Most people at risk of malaria live in areas of stable transmission…? is simply wrong. It is true that in many parts of the world malaria is termed “stable? because transmission remains relatively constant from year to year, the disease is endemic, the collective immunity is high, and epidemics are uncommon. However, in many other regions, the disease is endemic but “unstable? because annual transmission varies considerably, and the potential for epidemics is great. Climatic factors, particularly rainfall, are sometimes, but by no means always, relevant.15Again, “On the fringes of endemic zones, where transmission is limited by rainfall…there are strong seasonal patterns, and occasional major epidemics? is also wrong. In many regions, far from any “fringes?, malaria is endemic, stable, but highly seasonal. For example, in semi-arid regions of Mali, transmission is restricted to the rainy season, from July to September. The same 3 months constituted the transmission season for Plasmodium falciparum in Italy before it was eliminated.16 Paradoxically, in parts of the Sudan, rainfall is restricted to a month at most, but malaria is transmitted throughout the year. Female Anopheles gambiae survive drought and heat by resting in dwellings and other sheltered places.17 Blood feeding and transmission continue, but the mosquitoes do not develop eggs until the rains return. This phenomenon, termed gonotrophic dissociation, is remarkably similar to the winter survival strategy of Anopheles atroparvus, the principal vector of malaria in Holland until the mid 20th century.16By contrast, malaria is unstable in many regions that normally have abundant rainfall, and epidemics occur during periods of drought. An illustrative example is the catastrophic 1934–35 epidemic in Ceylon (now Sri Lanka), estimated to have killed 100·000 people.18 Worst hit was the south-western quadrant of the country, where average annual rainfall is greater than 250 cm, and malaria was endemic, but unstable and relatively infrequent. The dominant vector, Anopheles culicifacies, breeds along the banks of rivers and tends to be scarce in normal years. In the years 1928–33 there was abundant rainfall, river flow was high, A culicifacies was rare, and the human population was exceptionally malaria-free. However, after failure of two successive monsoons, the drying rivers produced colossal numbers of A culicifacies, and the resulting epidemic was exacerbated by the low collective immunity. In the drier parts of the island, where A culicifacies was dominant but transmission was more stable, immunity protected the population from the worst ravages of the disease.Hales and Woodward state that “the underlying problem? of the future “extension of seasonality? of malaria is “pollution of the atmosphere?, and call for rich countries to “recognise their obligations to the poorest by substantially reducing fossil-fuel consumption?. We understand public anxiety about climate change, but are concerned that many of these much-publicised predictions are ill informed and misleading. We urge those involved to pay closer attention to the complexities of this challenging subject. <br/
