RESUMEN
Algae biotechnology holds immense promise for revolutionizing the bioeconomy through the sustainable and scalable production of various bioproducts. However, their development has been hindered by the lack of advanced genetic tools. This study introduces a synthetic biology approach to develop such tools, focusing on the construction and testing of synthetic promoters. By analyzing conserved DNA motifs within the promoter regions of highly expressed genes across six different algal species, we identified cis-regulatory elements (CREs) associated with high transcriptional activity. Combining the algorithms POWRS, STREME, and PhyloGibbs, we predicted 1511 CREs and inserted them into a minimal synthetic promoter sequence in 1, 2, or 3 copies, resulting in 4533 distinct synthetic promoters. These promoters were evaluated in vivo for their capacity to drive the expression of a transgene in a high-throughput manner through next-generation sequencing post antibiotic selection and fluorescence-activated cell sorting. To validate our approach, we sequenced hundreds of transgenic lines showing high levels of GFP expression. Further, we individually tested 14 identified promoters, revealing substantial increases in GFP expressionâup to nine times higher than the baseline synthetic promoter, with five matching or even surpassing the performance of the native AR1 promoter. As a result of this study, we identified a catalog of CREs that can now be used to build superior synthetic algal promoters. More importantly, here we present a validated pipeline to generate building blocks for innovative synthetic genetic tools applicable to any algal species with a sequenced genome and transcriptome data set.
Asunto(s)
Biología Computacional , Regiones Promotoras Genéticas , Biología Sintética , Regiones Promotoras Genéticas/genética , Biología Computacional/métodos , Biología Sintética/métodos , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , AlgoritmosRESUMEN
The pharmacokinetics of moxifloxacin were investigated in buffalo calves following a single intravenous and intramuscular administration of moxifloxacin (5 mg kg(-1) body wt.). Moxifloxacin concentrations in plasma and urine were determined by microbiological assay. Pharmacokinetic analysis of disposition data indicated that intravenous administration data were best described by a two compartment open model, whereas intramuscular administration data were best described by a one compartment open model. Following intravenous administration, the elimination half life (t(1/2beta)), volume of distribution (Vd(area)) and total body clearance were 2.69+/-0.14 h, 1.43+/-0.08 L kg(-1) and 371.2+/-11.2 ml kg(-1)h(-1), respectively. Following intramuscular administration, the absorption half life (t(1/2ka)) was 0.83+/-0.20 h. The systemic bioavailability (F) of moxifloxacin in buffalo calves was 80.0+/-4.08%. Urinary excretion of moxifloxacin was less than 14% after 24h of administration of drug. In vitro binding of moxifloxacin to plasma proteins of buffalo calves was 28.4+/-3.77%. From the data of surrogate markers (AUC/MIC, C(max)/MIC), it was determined in the buffalo calves that when administered by intravenous or intramuscular route at 5 mg kg(-1), moxifloxacin is likely to be effective against bacterial isolates with MIC< or =0.1 microg ml(-1).
Asunto(s)
Antibacterianos/farmacocinética , Compuestos Aza/farmacocinética , Búfalos/metabolismo , Quinolinas/farmacocinética , Animales , Antibacterianos/administración & dosificación , Antibacterianos/sangre , Área Bajo la Curva , Compuestos Aza/administración & dosificación , Compuestos Aza/sangre , Disponibilidad Biológica , Búfalos/sangre , Fluoroquinolonas , Semivida , Masculino , Moxifloxacino , Quinolinas/administración & dosificación , Quinolinas/sangre , Distribución TisularRESUMEN
The costs of heart operations and the problems related to anticoagulation after prosthetic valve replacement are among the limitations faced by patients in nonindustrialized countries with mitral stenosis caused by chronic rheumatic heart disease. The young age at which these patients are seen also compels the surgeon to preserve the native valve. The least costly and optimal way to achieve this objective is by closed mitral valvotomy. After closed mitral valvotomy, mitral restenosis is commonly encountered. We report here our 10-year experience with operation on 113 consecutive patients with mitral restenosis. Closed transventricular revalvotomy was performed with Tubbs dilator in 105 of 113 patients. Mean age was 343 years, with a male to female ratio of 1:1.5. Most patients were in New York Heart Association functional classes III and IV (74.3% and 19.4%, respectively). Mean interval between first and second valvotomy was 9.4 years, Hospital mortality rate was 2.8%, trivial postoperative mitral regurgitation occurred in 16.1%, and moderately severe regurgitation occurred in 1.9%. Early postoperative systemic embolism occurred in 3.8% of the cases. Moderate to excellent symptomatic improvement was noted in 89.4% of the cases and poor results were seen in 10.2%. Late follow-up of 76 patients ranged from 2 to 10 years (mean 3.8 years), with 39.4% patients in New York Heart Association class I and 50% in class II. Close mitral revalvotomy is thus an economical, simple, and safe palliative procedure that carries good long-term results.