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Biomedical subjects

E Gulari

Publications and source records attributed to E Gulari.

6 recordsLinked to original sources
PubMed2004

Biphenyl and benzoate metabolism in a genomic context: outlining genome-wide metabolic networks in Burkholderia xenovorans LB400.

We designed and successfully implemented the use of in situ-synthesized 45-mer oligonucleotide DNA microarrays (XeoChips) for genome-wide expression profiling of Burkholderia xenovorans LB400, which is among the best aerobic polychlorinated biphenyl degraders known so far. We conducted differential gene expression profiling during exponential growth on succinate, benzoate, and biphenyl as sole carbon sources and investigated the transcriptome of early-stationary-phase cells grown on biphenyl. Based on these experiments, we outlined metabolic pathways and summarized other cellular functions in the organism relevant for biphenyl and benzoate degradation. All genes previously identified as being directly involved in biphenyl degradation were up-regulated when cells were grown on biphenyl compared to expression in succinate-grown cells. For benzoate degradation, however, genes for an aerobic coenzyme A activation pathway were up-regulated in biphenyl-grown cells, while the pathway for benzoate degradation via hydroxylation was up-regulated in benzoate-grown cells. The early-stationary-phase biphenyl-grown cells showed similar expression of biphenyl pathway genes, but a surprising up-regulation of C(1) metabolic pathway genes was observed. The microarray results were validated by quantitative reverse transcription PCR with a subset of genes of interest. The XeoChips showed a chip-to-chip variation of 13.9%, compared to the 21.6% variation for spotted oligonucleotide microarrays, which is less variation than that typically reported for PCR product microarrays.

Bacterial Proteins↗
PubMed2001

A flexible light-directed DNA chip synthesis gated by deprotection using solution photogenerated acids.

Oligonucleotide microarrays or oDNA chips are effective decoding and analytical tools for genomic sequences and are useful for a broad range of applications. Therefore, it is desirable to have synthesis methods of DNA chips that are highly flexible in sequence design and provide high quality and general adoptability. We report herein, DNA microarray synthesis based on a flexible biochip method. Our method simply uses photogenerated acid (PGA) in solution to trigger deprotection of the 5'-OH group in conventional nucleotide phosphoramidite monomers (i.e. PGA-gated deprotection), with the rest of the reactions in the synthesis cycle the same as those used for routine synthesis of oligonucleotides. The complete DNA chip synthesis process is accomplished on a regular DNA synthesizer that is coupled with a UV-VIS projection display unit for performing digital photolithography. Using this method, oDNA chips containing probes of newly discovered genes can be quickly and easily synthesized at high yields in a conventional laboratory setting. Furthermore, the PGA-gated chemistry should be applicable to microarray syntheses of a variety of combinatorial molecules, such as peptides and organic molecules.

DNA↗
PubMedDate not supplied

Digital light-directed synthesis. A microarray platform that permits rapid reaction optimization on a combinatorial basis.

Solution reactions using photogenerated reagents (Gao, X.; Yu, P.; LeProust, E.; Sonigo, L.; Pellois, J. P.; Zhang, H. J. Am. Chem. Soc. 1998, 120, 12698) are a potentially powerful means for combinatorial parallel synthesis of addressable molecular microarrays. In this report, we demonstrate that this chemistry permits combinatorial screening of reaction conditions on a microarray platform. Using this method of optimization and our reaction apparatus, efficient photogenerated acids and reaction conditions suitable for removal of the acid labile protection group on 5'-O of nucleotides are identified in a short period of time. The chemistry platform demonstrated opens new avenues for rapid, simultaneous investigation of multiple reactions using different reagents and reaction parameters directly on a solid support (e.g., a glass plate). The combinatorial screening method described may be extended to include general organic reactions employing photogenerated and conventional reagents as well as a microarray reaction device. This should be especially valuable for efficient synthesis of addressable organic compound libraries.

Chemistry, Organic↗
PubMedDate not supplied

Design engineering of a bioartificial renal tubule cell therapy device.

The use of a bioartificial renal tubule device composed of renal proximal tubule cells grown within a hollow fiber cartridge is a first step in engineering a bioartificial kidney to provide more complete replacement therapy of renal function than is available today. In this study, the feasibility of two designs for a tubule device were investigated: one with cells grown on microcarrier beads densely packed within the extracapillary space of a hollow fiber cartridge, and the other with cells grown as a confluent monolayer within the hollow fibers themselves. First, the oxygen requirements of porcine renal proximal tubule cells were determined, both attached to microcarriers and in suspension and compared to that of proximal tubule segments. The basal rate of cell respiration was found to be 2.29 +/- 0.53 nmol O2/10(6) cells/min for our cultured proximal tubule cells in suspension and no significant difference was seen with attached cells. Proximal tubule segments displayed significantly higher respiratory rates. Cells were also found to be responsive in the presence of mitochondrial inhibitors or uncouplers, and their respiratory rates remained constant, despite multiple passaging. The resultant cell oxygen consumption parameter was used in models describing oxygen concentration profiles within the two device configurations. From these models, it was found that cells within our proposed device designs could theoretically be sustained and remain viable, with respect to oxygen limitations. Finally, flow visualization studies were performed to assess fluid flow distribution and determine optimal device configuration and geometry to decrease areas of low or stagnant flow.

Animals↗