Results from high-throughput DNA cloning of Arabidopsis thaliana target genes using site-specific recombination

Thao, Sandy; Zhao, Qin; Kimball, Todd; Steffen, Eric; Blommel, Paul G.; Riters, Megan; Newman, Christopher; Fox, Brian G.; Wrobel, Russell L.

doi:10.1007/s10969-004-7148-4

Cited by 77 publications

(68 citation statements)

References 13 publications

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“…RT-PCR data were available for a subset of the genome containing 947 (3.5%) genes, which were chosen as part of a large protein expression study through the University of Wisconsin Center for Eukaryotic Structural Genomics (17). The RT-PCR cloning effort amplified 594 full-length cDNAs of 947 genes (Data Set 3, which is published as supporting information on the PNAS web site).…”

Section: Resultsmentioning

confidence: 99%

Identification of transcribed sequences in Arabidopsis thaliana by using high-resolution genome tiling arrays

Štolc

Samanta

Tongprasit

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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142

116

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Using a maskless photolithography method, we produced DNA oligonucleotide microarrays with probe sequences tiled throughout the genome of the plant Arabidopsis thaliana. RNA expression was determined for the complete nuclear, mitochondrial, and chloroplast genomes by tiling 5 million 36-mer probes. These probes were hybridized to labeled mRNA isolated from liquid grown T87 cells, an undifferentiated Arabidopsis cell culture line. Transcripts were detected from at least 60% of the nearly 26,330 annotated genes, which included 151 predicted genes that were not identified previously by a similar genome-wide hybridization study on four different cell lines. In comparison with previously published results with 25-mer tiling arrays produced by chromium masking-based photolithography technique, 36-mer oligonucleotide probes were found to be more useful in identifying intronexon boundaries. Using two-dimensional HPLC tandem mass spectrometry, a small-scale proteomic analysis was performed with the same cells. A large amount of strongly hybridizing RNA was found in regions ''antisense'' to known genes. Similarity of antisense activities between the 25-mer and 36-mer data sets suggests that it is a reproducible and inherent property of the experiments. Transcription activities were also detected for many of the intergenic regions and the small RNAs, including tRNA, small nuclear RNA, small nucleolar RNA, and microRNA. Expression of tRNAs correlates with genome-wide amino acid usage.higher plant ͉ transcriptome ͉ maskless array synthesizer

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Section: Resultsmentioning

confidence: 99%

Identification of transcribed sequences in Arabidopsis thaliana by using high-resolution genome tiling arrays

Štolc

Samanta

Tongprasit

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

142

116

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“…The genes encoding rASPA and hASPA were cloned, and selenomethionine-labeled proteins were expressed and purified following the standard Center for Eukary- otic Structural Genomics pipeline protocols for cloning (56), protein expression (57), protein purification (58), and overall bioinformatics management (59). Protein purification of heterologously expressed, nonglycosylated proteins from Escherichia coli did not involve refolding steps and resulted in 18.6 and 2.5 mg of recombinant rASPA and hASPA proteins, respectively.…”

Section: Methodsmentioning

confidence: 99%

Structure of aspartoacylase, the brain enzyme impaired in Canavan disease

Bitto

Bingman

Wesenberg

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

120

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Aspartoacylase catalyzes hydrolysis of N-acetyl-L-aspartate to aspartate and acetate in the vertebrate brain. Deficiency in this activity leads to spongiform degeneration of the white matter of the brain and is the established cause of Canavan disease, a fatal progressive leukodystrophy affecting young children. We present crystal structures of recombinant human and rat aspartoacylase refined to 2.8-and 1.8-Å resolution, respectively. The structures revealed that the N-terminal domain of aspartoacylase adopts a protein fold similar to that of zinc-dependent hydrolases related to carboxypeptidases A. The catalytic site of aspartoacylase shows close structural similarity to those of carboxypeptidases despite only 10 -13% sequence identity between these proteins. About 100 C-terminal residues of aspartoacylase form a globular domain with a two-stranded ␤-sheet linker that wraps around the N-terminal domain. The long channel leading to the active site is formed by the interface of the N-and C-terminal domains. The C-terminal domain is positioned in a way that prevents productive binding of polypetides in the active site. The structures revealed that residues 158 -164 may undergo a conformational change that results in opening and partial closing of the channel entrance. We hypothesize that the catalytic mechanism of aspartoacylase is closely analogous to that of carboxypeptidases. We identify residues involved in zinc coordination, and propose which residues may be involved in substrate binding and catalysis. The structures also provide a structural framework necessary for understanding the deleterious effects of many missense mutations of human aspartoacylase.N-acetyl-L-aspartate ͉ x-ray structure ͉ zinc-dependent hydrolase

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“…In Vitro Histone Peptide Pull-Down Assay cDNAs of PHD-finger domains of VIN3 and VIL1 were amplified by RT-PCR and cloned into pENTR-D-TOPO vector and transferred into a pVP13 expression vector (Thao et al, 2004) using LR reaction (Invitrogen). Recombinant proteins of VIN3 or VIL1 with His-tag were purified using Ni-NTA column (Qiagen).…”

Section: Western-blot Analysismentioning

confidence: 99%

The Binding Specificity of the PHD-Finger Domain of VIN3 Moderates Vernalization Response

Kim

Sung

2016

Plant Physiol.

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Results from high-throughput DNA cloning of Arabidopsis thaliana target genes using site-specific recombination

Cited by 77 publications

References 13 publications

Identification of transcribed sequences in Arabidopsis thaliana by using high-resolution genome tiling arrays

Identification of transcribed sequences in Arabidopsis thaliana by using high-resolution genome tiling arrays

Structure of aspartoacylase, the brain enzyme impaired in Canavan disease

The Binding Specificity of the PHD-Finger Domain of VIN3 Moderates Vernalization Response

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