Biomedical subjects

An-Ping Hsia

Publications and source records attributed to An-Ping Hsia.

6 recordsLinked to original sources

PubMed2005

Temperature gradient capillary electrophoresis (TGCE)--a tool for the high-throughput discovery and mapping of SNPs and IDPs.

Temperature gradient capillary electrophoresis (TGCE) can be used to distinguish heteroduplex from homoduplex DNA molecules and can thus be applied to the detection of various types of DNA polymorphisms. Unlike most single nucleotide polymorphism (SNP) detection technologies, TGCE can be used even in the absence of prior knowledge of the sequences of the underlying polymorphisms. TGCE is both sensitive and reliable in detecting SNPs, small InDel (insertion/deletion) polymorphisms (IDPs) and simple sequence repeats, and using this technique it is possible to detect a single SNP in amplicons of over 800 bp and 1-bp IDPs in amplicons of approximately 500 bp. Genotyping data obtained via TGCE are consistent with data obtained via gel-based detection technologies. For genetic mapping experiments, TGCE has a number of advantages over alternative heteroduplex-detection technologies such as celery endonuclease (CELI) and denaturing high-performance liquid chromatography (dHPLC). Multiplexing can increase TGCE's throughput to 12 markers on 94 recombinant inbreds per day. Given its ability to efficiently and reliably detect a variety of subtle DNA polymorphisms that occur at high frequency in genes, TGCE shows great promise for discovering polymorphisms and conducting genetic mapping and genotyping experiments.

Chromosome Mapping↗

PubMed2004

Types and frequencies of sequencing errors in methyl-filtered and high c0t maize genome survey sequences.

The Maize Genome Sequencing Consortium has deposited into GenBank more than 850,000 maize (Zea mays) genome survey sequences (GSSs) generated via two gene enrichment strategies, methylation filtration and high-C(0)t (HC) fractionation. These GSSs are a valuable resource for generating genome assemblies and the discovery of single nucleotide polymorphisms and nearly identical paralogs. Based on the rate of mismatches between 183 GSSs (105 methylation filtration + 78 HC) and 10 control genes, the rate of sequencing errors in these GSSs is 2.3 x 10(-3). As expected many of these errors were derived from insufficient vector trimming and base-calling errors. Surprisingly, however, some errors were due to cloning artifacts. These G.C to A.T transitions are restricted to HC clones; over 40% of HC clones contain at least one such artifact. Because it is not possible to distinguish the cloning artifacts from biologically relevant polymorphisms, HC sequences should be used with caution for the discovery of single nucleotide polymorphisms or paramorphisms. The average rate of sequencing errors was reduced 6-fold (to 3.6 x 10(-4)) by applying more stringent trimming parameters. This trimming resulted in the loss of only 11% of the bases (15,469/144,968). Due to redundancy among GSSs this more stringent trimming reduced coverage of promoters, exons, and introns by only 0%, 1%, and 4%, respectively. Hence, at the cost of a very modest loss of gene coverage, the quality of these maize GSSs can approach Bermuda standards, even prior to assembly.

DNA, Plant↗

PubMed2004

Picky: oligo microarray design for large genomes.

MOTIVATION: Many large genomes are getting sequenced nowadays. Biologists are eager to start microarray analysis taking advantage of all known genes of a species, but existing microarray design tools were very inefficient for large genomes. Also, many existing tools operate in a batch mode that does not assure best designs. RESULTS: Picky is an efficient oligo microarray design tool for large genomes. Picky integrates novel computer science techniques and the best known nearest-neighbor parameters to quickly identify sequence similarities and estimate their hybridization properties. Oligos designed by Picky are computationally optimized to guarantee the best specificity, sensitivity and uniformity under the given design constrains. Picky can be used to design arrays for whole genomes, or for only a subset of genes. The latter can still be screened against a whole genome to attain the same quality as a whole genome array, thereby permitting low budget, pathway-specific experiments to be conducted with large genomes. Picky is the fastest oligo array design tool currently available to the public, requiring only a few hours to process large gene sets from rice, maize or human.

Algorithms↗

PubMed2004

Mutation of a nitrate transporter, AtNRT1:4, results in a reduced petiole nitrate content and altered leaf development.

Unlike nitrate uptake of plant roots, less is known at the molecular level about how nitrate is distributed in various plant tissues. In the present study, characterization of the nitrate transporter, AtNRT1:4, revealed a special role of petiole in nitrate homeostasis. Electrophysiological studies using Xenopus oocytes showed that AtNRT1:4 was a low-affinity nitrate transporter. Whole-mount in situ hybridization and RT-PCR demonstrated that AtNRT1:4 was expressed in the leaf petiole. In the wild type, the leaf petiole had low nitrate reductase activity, but a high nitrate content, indicating that it is the storage site for nitrate, whereas, in the atnrt1:4 mutant, the petiole nitrate content was reduced to 50-64% of the wild-type level. Moreover, atnrt1:4 mutant leaves were wider than wild-type leaves. This study revealed a critical role of AtNRT1:4 in regulating leaf nitrate homeostasis, and the deficiency of AtNRT1:4 can alter leaf development.

Anion Transport Proteins↗

PubMed2004

The Etched1 gene of Zea mays (L.) encodes a zinc ribbon protein that belongs to the transcriptionally active chromosome (TAC) of plastids and is similar to the transcription factor TFIIS.

Etched1 (et1) is a pleiotropic, recessive mutation of maize that causes fissured and cracked mature kernels and virescent seedlings. Microscopic examinations of the et1 phenotype revealed an aberrant plastid development in mutant kernels and mutant leaves. Here, we report on the cloning of the et1 gene by transposon tagging, the localization of the gene product in chloroplasts, and its putative function in the plastid transcriptional apparatus. Several alleles of Mutator (Mu)-induced et1 mutants, the et1-reference (et1-R) mutant, and Et1 wild-type were cloned and analyzed at the molecular level. Northern analyses with wild-type plants revealed that Et1 transcripts are present in kernels, leaves, and other types of tissue, and no Et1 expression could be detected in the et1 mutants analyzed. The ET1 protein is imported by chloroplasts and has been immunologically detected in transcriptionally active chromosome (TAC) fractions derived from chloroplasts. Accordingly, the relative transcriptional activity of TAC fractions was significantly reduced in chloroplasts of et1-R plants. ET1 is the first zinc ribbon (ZR) protein shown to be targeted to plastids. With regard to its localization and its striking structural similarity to the eukaryotic transcription elongation factor TFIIS, it is feasible that ET1 functions in plastid transcription elongation by reactivation of arrested RNA polymerases.

Amino Acid Sequence↗

PubMed2003

Alternative transcription initiation sites and polyadenylation sites are recruited during Mu suppression at the rf2a locus of maize.

Even in the absence of excisional loss of the associated Mu transposons, some Mu-induced mutant alleles of maize can lose their capacity to condition a mutant phenotype. Three of five Mu-derived rf2a alleles are susceptible to such Mu suppression. The suppressible rf2a-m9437 allele has a novel Mu transposon insertion (Mu10) in its 5' untranslated region (UTR). The suppressible rf2a-m9390 allele has a Mu1 insertion in its 5' UTR. During suppression, alternative transcription initiation sites flanking the Mu1 transposon yield functional transcripts. The suppressible rf2a-m8110 allele has an rcy/Mu7 insertion in its 3' UTR. Suppression of this allele occurs via a previously unreported mechanism; sequences in the terminal inverted repeats of rcy/Mu7 function as alternative polyadenylation sites such that the suppressed rf2a-m8110 allele yields functional rf2a transcripts. No significant differences were observed in the nucleotide compositions of these alternative polyadenylation sites as compared with 94 other polyadenylation sites from maize genes.

Base Sequence↗