Isolation, characterization and mapping of temperature-sensitive mutants of mycobacteriophage I3

Eighteen temperature-sensitive mutants of mycobacteriophage I3 have been isolated and partially characterized. All the mutants were defective in vegetative replication. Based on temperature shift experiments with the temperature sensitive mutants, the thermosensitive phase of the phage development period has been characterized for each mutant. The genes have been mapped by recombination analysis. The early, continuous and middle genes seem to cluster on the genetic ma

Influence of formamide on the thermal stability of DNA

The utility of formamide in the denaturation and renaturation of DNA has been examined. The melting temperature of duplex DNA is lowered by 0.6°C per per cent formamide. The depression of melting temperature is independent of the GC content. Formamide also increases the width of the thermal transition. Upto 30%, it does not affect the rate of DNA reassociation

A rapid procedure for the isolation of spheroplasts from Mycobacterium smegmatis

Abstract is not available

Expression of drug resistance markers by spheroplasts ofMycobacterium smegmatis after fusion

Mycobacterial spheroplasts were prepared by treatment of the glycinesensitized cells with a combination of lipase and lysozyme. They were stable for several hours at room temperature but were lysed on treatment with 0.1% sodium dodecyl sulfate. The spheroplasts could be regenerated on a suitable medium. Fusion and regeneration of the spheroplasts were attempted using drug resistant mutant strains ofM. smegmalis. Recombinants were obtained from spheroplast fusion mediated by polyethylene glycol and dimethyl sulfoxide. Simultaneous expression of rccombinant properties was observed only after an initial lag in the isolated clones. This has been explained as due to “chromosome inactivation” in the fused product

A rapid procedure for the isolation of spheroplasts from Mycobacterium smegmatis

This article does not have an abstract

Influence of formamide on the thermal stability of DNA

The utility of formamide in the denaturation and renaturation of DNA has been examined. The mmelting temperature of duplex DNA is lowered by 0.6°C per per cent of formamide. The depression of melting temperature is independent of the GC content. Formamide also increases the width of the thermal transition. Upto 30% it does not affect the rate of DNA reassociation

Use of S<SUB>1</SUB> nuclease and formamide in combination for the reassociation studies on GC-rich DNA

The optimal parameters in the use of nuclease S<SUB>1</SUB> in DNA reassociation kinetics in the presence of formamide have been determined. The conditions are especially suitable for the study of DNA rich in mole percent GC. A 10-fold dilution of the reassociation samples leading to a decrease in both NaCl and formamide concentrations, consequently resulting in a lowering of T<SUB>m</SUB> by only 1.5°C, and the S<SUB>1</SUB> digestion at temperatures identical to the reassociation assay in order to retain the stability of the duplex, are two important aspects of this system. Under these conditions, the kinetics of reassociation followed the theoretically predicted pattern, while the earlier reported methods have shown lower values

Immune Responses and Immunosuppressive Strategies for Adeno-Associated Virus-Based Gene Therapy for Treatment of Central Nervous System Disorders: Current Knowledge and Approaches

Adeno-associated viruses (AAVs) are being increasingly used as gene therapy vectors in clinical studies especially targeting central nervous system (CNS) disorders. Correspondingly, host immune responses to the AAV capsid or the transgene-encoded protein have been observed in various clinical and preclinical studies. Such immune responses may adversely impact patients' health, prevent viral transduction, prevent repeated dosing strategies, eliminate transduced cells, and pose a significant barrier to the potential effectiveness of AAV gene therapy. Consequently, multiple immunomodulatory strategies have been used in attempts to limit immune-mediated responses to the vector, enable readministration of AAV gene therapy, prevent end-organ toxicity, and increase the duration of transgene-encoded protein expression. Herein we review the innate and adaptive immune responses that may occur during CNS-targeted AAV gene therapy as well as host- and treatment-specific factors that could impact the immune response. We also summarize the available preclinical and clinical data on immune responses specifically to CNS-targeted AAV gene therapy and discuss potential strategies for incorporating prophylactic immunosuppression regimens to circumvent adverse immune responses