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1.
EMBO J ; 43(15): 3240-3255, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38886582

RESUMO

Mutational patterns caused by APOBEC3 cytidine deaminase activity are evident throughout human cancer genomes. In particular, the APOBEC3A family member is a potent genotoxin that causes substantial DNA damage in experimental systems and human tumors. However, the mechanisms that ensure genome stability in cells with active APOBEC3A are unknown. Through an unbiased genome-wide screen, we define the Structural Maintenance of Chromosomes 5/6 (SMC5/6) complex as essential for cell viability when APOBEC3A is active. We observe an absence of APOBEC3A mutagenesis in human tumors with SMC5/6 dysfunction, consistent with synthetic lethality. Cancer cells depleted of SMC5/6 incur substantial genome damage from APOBEC3A activity during DNA replication. Further, APOBEC3A activity results in replication tract lengthening which is dependent on PrimPol, consistent with re-initiation of DNA synthesis downstream of APOBEC3A-induced lesions. Loss of SMC5/6 abrogates elongated replication tracts and increases DNA breaks upon APOBEC3A activity. Our findings indicate that replication fork lengthening reflects a DNA damage response to APOBEC3A activity that promotes genome stability in an SMC5/6-dependent manner. Therefore, SMC5/6 presents a potential therapeutic vulnerability in tumors with active APOBEC3A.


Assuntos
Proteínas de Ciclo Celular , Proteínas Cromossômicas não Histona , Citidina Desaminase , Dano ao DNA , Replicação do DNA , Humanos , Citidina Desaminase/metabolismo , Citidina Desaminase/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Cromossômicas não Histona/genética , Instabilidade Genômica , Linhagem Celular Tumoral , Proteínas
2.
J Adv Nurs ; 2024 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-38819604

RESUMO

AIM: To identify, synthesize and evaluate primary research on registered nurses' (RN) knowledge, attitudes and beliefs about sleep health and sleep health management of older adults living in residential aged care. DESIGN: Integrative review. DATA SOURCES: Medline, Embase and CINAHL databases from inception to September 2023. REVIEW METHODS: Databases were searched using a combination of key words, subject heading terms. All abstracts and full-text articles were screened by two researchers. Qualitative synthesis of the included articles was conducted. Inductive content analysis was used to identify themes and analyse data. RESULTS: A total of 923 abstracts were screened resulting in a final yield of 13 articles. Three themes were identified: (i) RN experience with sleep-disturbed residents, (ii) the emotional burden of sleep disturbances on RN and, (iii) organizational barriers to promoting resident's healthy sleep. Inappropriate administration of benzodiazepines and psychotropic drugs to manage residents' sleep disturbances was a major issue and lack of resources in residential aged care to facilitate sleep. There were concerns on nursing activity that disturbed residents' sleep and striking a balance between facilitating sleep and meeting managerial expectations was challenging. CONCLUSION: This review identified that nurses' decision-making has an integral role in the management of sleep health in residents in aged care. Whilst evidence-based guidelines for managing sleep in residential aged care are available, there is a lack of translation to practice. Understanding RN perspectives is critical to improving sleep health models of care in residential aged care. IMPACT: This review found that RN are attuned to the implications of sleep disturbance in residential aged care but are constrained by current sleep health models of care. PATIENT OR PUBLIC CONTRIBUTION: Not applicable.

3.
J Phys Chem C Nanomater Interfaces ; 128(29): 12194-12205, 2024 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-39081556

RESUMO

We present the vibrational properties and phonon dispersion for quasi-2D hybrid organic-inorganic perovskites (BA)2CsPb2I7, (HA)2CsPb2I7, (BA)2(MA)Pb2I7, and (HA)2(MA)Pb2I7 calculated from first principles. Given the highly complex nature of these compounds, we first perform careful benchmarking and convergence testing to identify suitable parameters to describe their structural features and vibrational properties. We find that the inclusion of van der Waals corrections on top of generalized gradient approximation (GGA) exchange-correlation functionals provides the best agreement for the equilibrium structure relative to experimental data. We also investigate the impact of the molecular orientation on the equilibrium structure of these layered perovskite systems. Our results suggest ground state ferroelectric alignment of molecular dipoles in the out-of-plane direction is unlikely and support the assignment of the centrosymmetric space group for the low-temperature phase of (HA)2(MA)Pb2I7. Finally, we compute vibrational properties under the harmonic approximation. We find that stringent energy cut-offs are required to obtain well-converged phonon properties, and once converged, the harmonic approximation can capture key physics for such a large, hybrid inorganic-organic system with vastly different atom types, masses, and interatomic interactions. We discuss the obtained phonon modes and dispersion behavior in the context of known properties for bulk 3D perovskites and ligand molecular crystals. While many vibrational properties are inherited from the parent systems, we also observe unique coupled vibrations that cannot be associated with vibrations of the pure constituent perovskite and ligand subphases. Energy dispersion of the low energy phonon branches primarily occurs in the in-plane direction and within the perovskite subphase and arises from bending and breathing modes of the equatorial Pb-I network within the perovskite octahedral plane. The analysis herein provides the foundation for future investigations on this class of materials, such as exciton-phonon coupling, phase transitions, and general temperature-dependent properties.

4.
JAMA Netw Open ; 7(4): e248889, 2024 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-38662368

RESUMO

Importance: With older drivers representing the fastest growing segment of the driver population and dementia prevalence increasing with age, policymakers face the challenge of balancing road safety and mobility of older adults. In states that require reporting a dementia diagnosis to the Department of Motor Vehicles (DMV), individuals with dementia may be reluctant to disclose symptoms of cognitive decline, and clinicians may be reluctant to probe for those symptoms, which may be associated with missed or delayed diagnoses. Objective: To assess whether DMV reporting policies for drivers with dementia are associated with primary care clinicians' underdiagnosing dementia. Design, Setting, and Participants: This cross-sectional study used data from the 100% Medicare fee-for-service program and the Medicare Advantage plans from 2017 to 2019 on 223 036 primary care clinicians with at least 25 Medicare patients. Statistical analysis was performed from July to October 2023. Exposures: State DMV reporting policies for drivers with dementia. Main Outcomes and Measures: The main outcome was a binary variable indicating whether the clinician underdiagnosed dementia or not. Each clinician's expected number of dementia cases was estimated using a predictive model based on patient characteristics. Comparing the estimation with observed dementia diagnoses identified clinicians who underdiagnosed dementia vs those who did not, after accounting for sampling errors. Results: Four states have clinician reporting mandates, 14 have mandates requiring drivers to self-report dementia diagnoses, and 32 states and the District of Columbia do not have explicit requirements. Among primary care clinicians in states with clinician reporting mandates (n = 35 620), 51.4% were female, 91.9% worked in a metropolitan area, and 19.9% of the patient panel were beneficiaries dually eligible for Medicare and Medicaid. Among primary care clinicians in states with patient self-reporting mandates (n = 57 548), 55.7% were female, 83.1% worked in a metropolitan area, and 15.4% of the patient panel were dually eligible for Medicare and Medicaid. Among clinicians in states without mandates, 55.7% were female, 83.0% worked in a metropolitan area, and 14.6% of the patient panel were dually eligible for Medicare and Medicaid. Clinicians in states with clinician reporting mandates had an adjusted 12.4% (95% CI, 10.5%-14.2%) probability of underdiagnosing dementia compared with 7.8% (95% CI, 6.9%-8.7%) in states with self-reporting and 7.7% (95% CI, 6.9%-8.4%) in states with no mandates, an approximately 4-percentage point difference (P < .001). Conclusions and Relevance: Results of this cross-sectional study of primary care clinicians suggest that mandatory DMV policies for clinicians to report patients with dementia may be associated with a higher risk of missed or delayed dementia diagnoses. Future research is needed to better understand the unintended consequences and the risk-benefit tradeoffs of these policies.


Assuntos
Demência , Medicare , Humanos , Demência/diagnóstico , Demência/epidemiologia , Estudos Transversais , Estados Unidos , Feminino , Masculino , Idoso , Medicare/estatística & dados numéricos , Condução de Veículo/legislação & jurisprudência , Condução de Veículo/estatística & dados numéricos , Notificação de Abuso , Idoso de 80 Anos ou mais
5.
Cureus ; 16(5): e61321, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38947683

RESUMO

Carcinoid syndrome is a rare condition resulting from neuroendocrine tumors (NETs) that secrete vasoactive substances like serotonin. This report describes the case of a 61-year-old man with a history of chronic obstructive pulmonary disease (COPD) and hypertension who presented with new-onset angioedema, loss of consciousness, and a fall. He had been treated for COPD exacerbations during ER visits without improvement and was unaware of a prior mesenteric carcinoid tumor diagnosis from 2012. The next emergency evaluation revealed significant airway and facial edema necessitating intubation. Imaging and biopsy identified a well-differentiated grade 1 NET with extensive liver metastases. Laboratory tests showed elevated levels of serum serotonin, chromogranin A, and 24-hour urine 5-hydroxyindoleacetic acid (5-HIAA). Post-discharge, a PET scan confirmed metastatic lesions primarily in the liver and small bowel, with an unresectable mesenteric mass. The patient was treated with lanreotide and became symptom-free. This case underscores the need to consider carcinoid syndrome in patients with COPD presenting with unexplained respiratory symptoms, as timely diagnosis and treatment can significantly enhance patient outcomes.

6.
bioRxiv ; 2024 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-38746251

RESUMO

Humans effortlessly use vision to plan and guide navigation through the local environment, or "scene". A network of three cortical regions responds selectively to visual scene information, including the occipital (OPA), parahippocampal (PPA), and medial place areas (MPA) - but how this network supports visually-guided navigation is unclear. Recent evidence suggests that one region in particular, the OPA, supports visual representations for navigation, while PPA and MPA support other aspects of scene processing. However, most previous studies tested only static scene images, which lack the dynamic experience of navigating through scenes. We used dynamic movie stimuli to test whether OPA, PPA, and MPA represent two critical kinds of navigationally-relevant information: navigational affordances (e.g., can I walk to the left, right, or both?) and ego-motion (e.g., am I walking forward or backward? turning left or right?). We found that OPA is sensitive to both affordances and ego-motion, as well as the conflict between these cues - e.g., turning toward versus away from an open doorway. These effects were significantly weaker or absent in PPA and MPA. Responses in OPA were also dissociable from those in early visual cortex, consistent with the idea that OPA responses are not merely explained by lower-level visual features. OPA responses to affordances and ego-motion were stronger in the contralateral than ipsilateral visual field, suggesting that OPA encodes navigationally relevant information within an egocentric reference frame. Taken together, these results support the hypothesis that OPA contains visual representations that are useful for planning and guiding navigation through scenes.

7.
bioRxiv ; 2024 Aug 06.
Artigo em Inglês | MEDLINE | ID: mdl-39149314

RESUMO

Generative pretrained models represent a significant advancement in natural language processing and computer vision, which can generate coherent and contextually relevant content based on the pre-training on large general datasets and fine-tune for specific tasks. Building foundation models using large scale omic data is promising to decode and understand the complex signaling language patterns within cells. Different from existing foundation models of omic data, we build a foundation model, mosGraphGPT , for multi-omic signaling (mos) graphs, in which the multi-omic data was integrated and interpreted using a multi-level signaling graph. The model was pretrained using multi-omic data of cancers in The Cancer Genome Atlas (TCGA), and fine-turned for multi-omic data of Alzheimer's Disease (AD). The experimental evaluation results showed that the model can not only improve the disease classification accuracy, but also is interpretable by uncovering disease targets and signaling interactions. And the model code are uploaded via GitHub with link: https://github.com/mosGraph/mosGraphGPT.

8.
bioRxiv ; 2024 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-39026868

RESUMO

Neurodegenerative diseases are often characterized by mitochondrial dysfunction. In Alzheimer's disease, abnormal tau phosphorylation disrupts mitophagy, a quality control process through which damaged organelles are selectively removed from the mitochondrial network. The precise mechanism through which this occurs remains unclear. Previously, we showed that tau which has been mutated at Thr-231 to glutamic acid to mimic an Alzheimer's-relevant phospho-epitope expressed early in disease selectively inhibits oxidative stress-induced mitophagy in C. elegans. Here, we use immortalized mouse hippocampal neuronal cell lines to extend that result into mammalian cells. Specifically, we show that phosphomimetic tau at Ser-396/404 (EC) or Thr-231/Ser-235 (EM) partly inhibits mitophagy induction by paraquat, a potent inducer of mitochondrial oxidative stress. Moreover, a combination of immunologic and biochemical approaches demonstrates that the levels of the mitophagy receptor FKBP8, significantly decrease in response to paraquat in cells expressing EC or EM tau mutants, but not in cells expressing wildtype tau. In contrast, paraquat treatment results in a decrease in the levels of the mitophagy receptors FUNDC1 and BNIP3 in the presence of both wildtype tau and the tau mutants. Interestingly, FKBP8 is normally trafficked to the endoplasmic reticulum during oxidative stress induced mitophagy, and our results support a model where this trafficking is impacted by disease-relevant tau, perhaps through a direct interaction. We provide new insights into the molecular mechanisms underlying tau pathology in Alzheimer's disease and highlight FKBP8 receptor as a potential target for mitigating mitochondrial dysfunction in neurodegenerative diseases.

9.
Nat Commun ; 15(1): 3156, 2024 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-38605017

RESUMO

Modulating brain oscillations has strong therapeutic potential. Interventions that both non-invasively modulate deep brain structures and are practical for chronic daily home use are desirable for a variety of therapeutic applications. Repetitive audio-visual stimulation, or sensory flicker, is an accessible approach that modulates hippocampus in mice, but its effects in humans are poorly defined. We therefore quantified the neurophysiological effects of flicker with high spatiotemporal resolution in patients with focal epilepsy who underwent intracranial seizure monitoring. In this interventional trial (NCT04188834) with a cross-over design, subjects underwent different frequencies of flicker stimulation in the same recording session with the effect of sensory flicker exposure on local field potential (LFP) power and interictal epileptiform discharges (IEDs) as primary and secondary outcomes, respectively. Flicker focally modulated local field potentials in expected canonical sensory cortices but also in the medial temporal lobe and prefrontal cortex, likely via resonance of stimulated long-range circuits. Moreover, flicker decreased interictal epileptiform discharges, a pathological biomarker of epilepsy and degenerative diseases, most strongly in regions where potentials were flicker-modulated, especially the visual cortex and medial temporal lobe. This trial met the scientific goal and is now closed. Our findings reveal how multi-sensory stimulation may modulate cortical structures to mitigate pathological activity in humans.


Assuntos
Epilepsias Parciais , Epilepsia , Humanos , Encéfalo , Eletroencefalografia , Lobo Temporal , Estudos Cross-Over
10.
iScience ; 27(3): 109083, 2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38361627

RESUMO

Exercise mediates tissue metabolic function through direct and indirect adaptations to acylcarnitine (AC) metabolism, but the exact mechanisms are unclear. We found that circulating medium-chain acylcarnitines (AC) (C12-C16) are lower in active/endurance trained human subjects compared to sedentary controls, and this is correlated with elevated cardiorespiratory fitness and reduced adiposity. In mice, exercise reduced serum AC and increased liver AC, and this was accompanied by a marked increase in expression of genes involved in hepatic AC metabolism and mitochondrial ß-oxidation. Primary hepatocytes from high-fat fed, exercise trained mice had increased basal respiration compared to hepatocytes from high-fat fed sedentary mice, which may be attributed to increased Ca2+ cycling and lipid uptake into mitochondria. The addition of specific medium- and long-chain AC to sedentary hepatocytes increased mitochondrial respiration, mirroring the exercise phenotype. These data indicate that AC redistribution is an exercise-induced mechanism to improve hepatic function and metabolism.

11.
NPJ Precis Oncol ; 8(1): 121, 2024 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-38806586

RESUMO

Cerebrospinal fluid tumor-derived DNA (CSF-tDNA) analysis is a promising approach for monitoring the neoplastic processes of the central nervous system. We applied a lung cancer-specific sequencing panel (CAPP-Seq) to 81 CSF, blood, and tissue samples from 24 lung cancer patients who underwent lumbar puncture (LP) for suspected leptomeningeal disease (LMD). A subset of the cohort (N = 12) participated in a prospective trial of osimertinib for refractory LMD in which serial LPs were performed before and during treatment. CSF-tDNA variant allele fractions (VAFs) were significantly higher than plasma circulating tumor DNA (ctDNA) VAFs (median CSF-tDNA, 32.7%; median plasma ctDNA, 1.8%; P < 0.0001). Concentrations of tumor DNA in CSF and plasma were positively correlated (Spearman's ρ, 0.45; P = 0.03). For LMD diagnosis, cytology was 81.8% sensitive and CSF-tDNA was 91.7% sensitive. CSF-tDNA was also strongly prognostic for overall survival (HR = 7.1; P = 0.02). Among patients with progression on targeted therapy, resistance mutations, such as EGFR T790M and MET amplification, were common in peripheral blood but were rare in time-matched CSF, indicating differences in resistance mechanisms based on the anatomic compartment. In the osimertinib cohort, patients with CNS progression had increased CSF-tDNA VAFs at follow-up LP. Post-osimertinib CSF-tDNA VAF was strongly prognostic for CNS progression (HR = 6.2, P = 0.009). Detection of CSF-tDNA in lung cancer patients with suspected LMD is feasible and may have clinical utility. CSF-tDNA improves the sensitivity of LMD diagnosis, enables improved prognostication, and drives therapeutic strategies that account for spatial heterogeneity in resistance mechanisms.

12.
JCO Oncol Pract ; : OP2400021, 2024 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-39028923

RESUMO

PURPOSE: The time required for in-clinic drug administration can substantially affect breast cancer patients' quality of life. Subcutaneous (SC) drug administration, as opposed to intravenous (IV), may reduce this time commitment. This study sought to estimate the difference in time burden between IV and SC administration of trastuzumab and pertuzumab (HP). METHODS: We prospectively enrolled a subcohort of patients participating in the ADEPT trial (ClinicalTrials.gov identifier: NCT04569747, investigating adjuvant HP plus endocrine therapy for stage I human epidermal growth factor receptor 2-positive breast cancer) to this single-arm crossover time and motion substudy. Patients received two cycles of IV HP followed by two cycles of SC HP. During each cycle, time points in drug preparation and administration were captured. The primary end point was total patient time in the treatment chair. Additional end points included total patient treatment experience time and total pharmacy workflow time. A sample size of 22 patients was estimated to provide 90.7% power with two-sided alpha .05 to detect a difference of 70 minutes in the primary end point by treatment arm (IV v SC). RESULTS: Twenty-two patients were enrolled. The mean total patient time in the treatment chair was 61.8 minutes shorter with SC versus IV HP (22.5 v 84.3 minutes; P < .0001). The mean total patient treatment experience time (incorporating time spent waiting for treatment initiation and time spent in the treatment chair) was 81.8 minutes shorter for SC administration (96 v 177.8 minutes; P < .0001). The pharmacy workflow time was 78.2 minutes shorter for SC versus IV formulation (41 v 119.2 minutes; P < .0001). CONCLUSION: SC administration of HP shortened patient time burden by approximately 1 hour. SC drug administration can facilitate faster workflows for health care professionals and improve patients' breast cancer treatment experience.

13.
Neurobiol Lang (Camb) ; 4(4): 575-610, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-38144236

RESUMO

Much of the language we encounter in our everyday lives comes in the form of conversation, yet the majority of research on the neural basis of language comprehension has used input from only one speaker at a time. Twenty adults were scanned while passively observing audiovisual conversations using functional magnetic resonance imaging. In a block-design task, participants watched 20 s videos of puppets speaking either to another puppet (the dialogue condition) or directly to the viewer (the monologue condition), while the audio was either comprehensible (played forward) or incomprehensible (played backward). Individually functionally localized left-hemisphere language regions responded more to comprehensible than incomprehensible speech but did not respond differently to dialogue than monologue. In a second task, participants watched videos (1-3 min each) of two puppets conversing with each other, in which one puppet was comprehensible while the other's speech was reversed. All participants saw the same visual input but were randomly assigned which character's speech was comprehensible. In left-hemisphere cortical language regions, the time course of activity was correlated only among participants who heard the same character speaking comprehensibly, despite identical visual input across all participants. For comparison, some individually localized theory of mind regions and right-hemisphere homologues of language regions responded more to dialogue than monologue in the first task, and in the second task, activity in some regions was correlated across all participants regardless of which character was speaking comprehensibly. Together, these results suggest that canonical left-hemisphere cortical language regions are not sensitive to differences between observed dialogue and monologue.

14.
Elife ; 122023 Dec 21.
Artigo em Inglês | MEDLINE | ID: mdl-38126343

RESUMO

Yes-associated protein (YAP), the downstream effector of the evolutionarily conserved Hippo pathway, promotes cellular proliferation and coordinates certain regenerative responses in mammals. Small molecule activators of YAP may, therefore, display therapeutic utility in treating disease states involving insufficient proliferative repair. From a high-throughput chemical screen of the comprehensive drug repurposing library ReFRAME, here we report the identification of SM04690, a clinical stage inhibitor of CLK2, as a potent activator of YAP-driven transcriptional activity in cells. CLK2 inhibition promotes alternative splicing of the Hippo pathway protein AMOTL2, producing an exon-skipped gene product that can no longer associate with membrane-bound proteins, resulting in decreased phosphorylation and membrane localization of YAP. This study reveals a novel mechanism by which pharmacological perturbation of alternative splicing inactivates the Hippo pathway and promotes YAP-dependent cellular growth.


Assuntos
Proteínas Serina-Treonina Quinases , Transdução de Sinais , Animais , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/fisiologia , Fatores de Transcrição/metabolismo , Processamento Alternativo , Proteínas de Sinalização YAP , Proteínas de Membrana/metabolismo , Mamíferos/metabolismo
15.
MDM Policy Pract ; 8(1): 23814683231178033, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-38178866

RESUMO

Introduction: Decision aids (DAs) are helpful instruments used to support shared decision making (SDM). Patients with atrial fibrillation (AF) face complex decisions regarding stroke prevention strategies. While a few DAs have been made for AF stroke prevention, an encounter DA (EDA) and patient DA (PDA) have not been created to be used in conjunction with each other before. Design: Using iterative user-centered design, we developed 2 DAs for anticoagulation choice and stroke prevention in AF. Prototypes were created, and we elicited feedback from patients and experts via observations of encounters, usability testing, and semistructured interviews. Results: User testing was done with 33 experts (in AF and SDM) and 51 patients from 6 institutions. The EDA and PDA underwent 1 and 4 major iterations, respectively. Major differences between the DAs included AF pathophysiology and a preparation to meet with the clinician in the PDA as well as different language throughout. Content areas included personalized stroke risk, differences between anticoagulants, and risks of bleeding. Based on user feedback, developers 1) addressed feelings of isolation with AF, 2) improved navigation options, 3) modified content and flow for users new to AF and those experienced with AF, 4) updated stroke risk pictographs, and 5) added structure to the preparation for decision making in the PDA. Limitations: These DAs focus only on anticoagulation for stroke prevention and are online, which may limit participation for those less comfortable with technology. Conclusions: Designing complementary DAs for use in tandem or separately is a new method to support SDM between patients and clinicians. Extensive user testing is essential to creating high-quality tools that best meet the needs of those using them. Highlights: First-time complementary encounter and patient decision aids have been designed to work together or separately.User feedback led to greater structure and different experiences for patients naïve or experienced with anticoagulants in patient decision aids.Online tools allow for easier dissemination, use in telehealth visits, and updating as new evidence comes out.

17.
Electron. j. biotechnol ; 15(4): 9-9, July 2012. ilus, tab
Artigo em Inglês | LILACS | ID: lil-646959

RESUMO

In the present study, a novel plant transformation system for Doritaenopsis and Phalaenopsis has been developed. The pollen-mediated activation tagging system was established by artificial pollination. The pollens, co-cultured with Agrobacterium tumefaciens strain EHA105 harbouring an activation tagging vector (pTAG-8), were used for pollination. In order to optimize the transformation efficiency, several factors (concentration of A. tumefaciens, concentration of acetosyringone during co-cultivation and the duration of co-cultivation) known to influence Agrobacterium-mediated DNA transfer were examined. A concentration of 0.5-1 x 10(8) CFU/ml for A. tumefaciens, 0.1 mM acetosyringone, and 6 hrs of co-culture period were found to be the optimal condition for high transformation efficiency. Integration of T-DNA into the genome of putative transgenic plants was confirmed by PCR and DNA blot analyses. Single copy of the transgene was observed in all transgenic plants analyzed. Most of the transgenic plants had a morphologically normal phenotype and the overall capsule formation efficiency was similar to control plant. Our results showed a new approach of genetic transformation in orchids and this method can be employed for genetic improvement of the orchids.


Assuntos
Agrobacterium tumefaciens , Orchidaceae/genética , Polinização , Elementos de DNA Transponíveis/genética , Reação em Cadeia da Polimerase , Transformação Genética
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