Biomedical subjects

Ming-Cheng Luo

Publications and source records attributed to Ming-Cheng Luo.

9 recordsLinked to original sources

PubMed2006

Efficacy of clone fingerprinting methodologies.

With the development of new high-information content fingerprinting techniques for constructing BAC-based physical maps, physical map construction is accelerating and it is important to determine which methodologies work best. In a recent publication (Z. Xu et al., 2004, Genomics 84:941-951), Xu et al. evaluated five different techniques (one agarose-based and four using multiple enzymes) and concluded that a two-enzyme technique was superior. In addition, they found that no benefit was gained from fingerprinting more than 10x coverage. In this paper we report our own extensive simulation results, which lead to contrasting conclusions. Our data indicate that the five-enzyme method known as SNaPshot is the most effective and that the assembly can in fact be significantly improved with greater than 10x coverage.

Arabidopsis↗

PubMed2006

GenoProfiler: batch processing of high-throughput capillary fingerprinting data.

UNLABELLED: High-throughput content fingerprinting techniques employing capillary electrophoresis place new demands on the editing of fingerprint files for the downstream contig assembly program, FPC. A cross-platform software application, GenoProfiler, was developed for automated editing of sized fingerprinting profiles generated by the ABI Genetic Analyzers. The batch-processing module extracts the sized fragment information directly from the ABI raw trace files, or from data files exported from GeneMapper or other size calling software, removes the background noise and undesired fragments, and generates fragment size files compatible with the FPC software. AVAILABILITY: http://wheat.pw.usda.gov/PhysicalMapping/

Algorithms↗

PubMed2006

Molecular characterization of a diagnostic DNA marker for domesticated tetraploid wheat provides evidence for gene flow from wild tetraploid wheat to hexaploid wheat.

All forms of domesticated tetraploid wheat (Triticum turgidum, genomes AABB) are nearly monomorphic for restriction fragment length polymorphism (RFLP) haplotype a at the Xpsr920 locus on chromosome 4A (Xpsr920-A1a), and wild tetraploid wheat is monomorphic for haplotype b. The Xpsr920-A1a/b dimorphism provides a molecular marker for domesticated and wild tetraploid wheat, respectively. Hexaploid wheat (Triticum aestivum, genomes AABBDD) is polymorphic for the 2 haplotypes. Bacterial artificial chromosome (BAC) clones hybridizing with PSR920 were isolated from Triticum urartu (genomes AA), Triticum monococcum (genomes AmAm), and T. turgidum ssp. durum (genomes AABB) and sequenced. PSR920 is a fragment of a putative ATP binding cassette (ABC) transporter gene (designated ABCT-1). The wheat ABCT-1 gene is more similar to the T. urartu gene than to the T. monococcum gene and diverged from the T. urartu gene about 0.7 MYA. The comparison of the sequence of the wheat A genome BAC clone with that of the T. urartu BAC clone provides the first insight into the microsynteny of the wheat A genome with that of T. urartu. Within 103 kb of orthologous intergenic space, 37 kb of new DNA has been inserted and 36 kb deleted leaving 49.7% of the region syntenic between the clones. The nucleotide substitution rate in the syntenic intergenic space has been 1.6 x 10(-8) nt(-1) year(-1), which is, respectively, 4 and 3 times as great as nucleotide substitution rates in the introns and the third codon positions of the juxtaposed gene. The RFLP is caused by a miniature inverted transposable element (MITE) insertion into intron 18 of the ABCT-A1 gene. Polymerase chain reaction primers were developed for the amplification of the MITE insertion site and its sequencing. The T. aestivum ABCT-A1a haplotype is identical to the haplotype of domesticated tetraploid wheat, and the ABCT-A1b haplotype is identical to that of wild tetraploid wheat. This finding shows for the first time that wild tetraploid wheat participated in the evolution of hexaploid wheat. A cline of the 2 haplotype frequencies exists across Euro-Asia in T. aestivum. It is suggested that T. aestivum in eastern Asia conserved the gene pool of the original T. aestivum more than wheat elsewhere.

Chromosomes, Plant↗

PubMed2006

Construction and characterization of two BAC libraries from Brachypodium distachyon, a new model for grass genomics.

Brachypodium is well suited as a model system for temperate grasses because of its compact genome and a range of biological features. In an effort to develop resources for genome research in this emerging model species, we constructed 2 bacterial artificial chromosome (BAC) libraries from an inbred diploid Brachypodium distachyon line, Bd21, using restriction enzymes HindIII and BamHI. A total of 73,728 clones (36,864 per BAC library) were picked and arrayed in 192,384-well plates. The average insert size for the BamHI and HindIII libraries is estimated to be 100 and 105 kb, respectively, and inserts of chloroplast origin account for 4.4% and 2.4%, respectively. The libraries individually represent 9.4- and 9.9-fold haploid genome equivalents with combined 19.3-fold genome coverage, based on a genome size of 355 Mb reported for the diploid Brachypodium, implying a 99.99% probability that any given specific sequence will be present in each library. Hybridization of the libraries with 8 starch biosynthesis genes was used to empirically evaluate this theoretical genome coverage; the frequency at which these genes were present in the library clones gave an estimated coverage of 11.6- and 19.6-fold genome equivalents. To obtain a first view of the sequence composition of the Brachypodium genome, 2185 BAC end sequences (BES) representing 1.3 Mb of random genomic sequence were compared with the NCBI GenBank database and the GIRI repeat database. Using a cutoff expectation value of E<10-10, only 3.3% of the BESs showed similarity to repetitive sequences in the existing database, whereas 40.0% had matches to the sequences in the EST database, suggesting that a considerable portion of the Brachypodium genome is likely transcribed. When the BESs were compared with individual EST databases, more matches hit wheat than maize, although their EST collections are of a similar size, further supporting the close relationship between Brachypodium and the Triticeae. Moreover, 122 BESs have significant matches to wheat ESTs mapped to individual chromosome bin positions. These BACs represent colinear regions containing the mapped wheat ESTs and would be useful in identifying additional markers for specific wheat chromosome regions.

Chromosomes, Artificial, Bacterial↗

PubMed2005

Comparative genetic maps reveal extreme crossover localization in the Aegilops speltoides chromosomes.

A total of 137 loci were mapped in Aegilops speltoides, the closest extant relative of the wheat B genome, using two F(2) mapping populations and a set of wheat-Ae. speltoides disomic addition (DA) lines. Comparisons of Ae. speltoides genetic maps with those of Triticum monococcum indicated that Ae. speltoides conserved the gross chromosome structure observed across the tribe Triticeae. A putative inversion involving the short arm of chromosome 2 was detected in Ae. speltoides. A translocation between chromosomes 2 and 6, present in the wheat B genome, was absent. The ligustica/aucheri spike dimorphism behaved as allelic variation at a single locus, which was mapped in the centromeric region of chromosome 3. The genetic length of each chromosome arm was about 50 cM, irrespective of its physical length. Compared to T. monococcum genetic maps, recombination was virtually eliminated from the proximal 50-100 cM and was localized in short distal regions, which were often expanded compared to the T. monococcum maps. The wheat B genome and the genome of Ae. longissima, a close relative of Ae. speltoides, do not show the extreme localization of crossovers observed in Ae. speltoides.

Chromosome Mapping↗

PubMed2005

The R1 resistance gene cluster contains three groups of independently evolving, type I R1 homologues and shows substantial structural variation among haplotypes of Solanum demissum.

Cultivated and wild potatoes contain a major disease-resistance cluster on the short arm of chromosome V, including the R1 resistance (R) gene against potato late blight. To explore the functional and evolutionary significance of clustering in the generation of novel disease-resistance genes, we constructed three approximately 1 Mb physical maps in the R1 gene region, one for each of the three genomes (haplotypes) of allohexaploid Solanum demissum, the wild potato progenitor of the R1 locus. Totals of 691, 919 and 559 kb were sequenced for each haplotype, and three distinct resistance-gene families were identified, one homologous to the potato R1 gene and two others homologous to either the Prf or the Bs4 R-gene of tomato. The regions with R1 homologues are highly divergent among the three haplotypes, in contrast to the conserved flanking non-resistance gene regions. The R1 locus shows dramatic variation in overall length and R1 homologue number among the three haplotypes. Sequence comparisons of the R1 homologues show that they form three distinct clades in a distance tree. Frequent sequence exchanges were detected among R1 homologues within each clade, but not among those in different clades. These frequent sequence exchanges homogenized the intron sequences of homologues within each clade, but did not homogenize the coding sequences. Our results suggest that the R1 homologues represent three independent groups of fast-evolving type I resistance genes, characterized by chimeric structures resulting from frequent sequence exchanges among group members. Such genes were first identified among clustered RGC2 genes in lettuce, where they were distinguished from slow-evolving type II R-genes. Our findings at the R1 locus in S. demissum may indicate that a common or similar mechanism underlies the previously reported differentiation of type I and type II R-genes and the differentiation of type I R-genes into distinct groups, identified here.

Chromosomes, Artificial, Bacterial↗

PubMed2004

Sequence polymorphism in polyploid wheat and their d-genome diploid ancestor.

Sequencing was used to investigate the origin of the D genome of the allopolyploid species Triticum aestivum and Aegilops cylindrica. A 247-bp region of the wheat D-genome Xwye838 locus, encoding ADP-glucopyrophosphorylase, and a 326-bp region of the wheat D-genome Gss locus, encoding granule-bound starch synthase, were sequenced in a total 564 lines of hexaploid wheat (T. aestivum, genome AABBDD) involving all its subspecies and 203 lines of Aegilops tauschii, the diploid source of the wheat D genome. In Ae. tauschii, two SNP variants were detected at the Xwye838 locus and 11 haplotypes at the Gss locus. Two haplotypes with contrasting frequencies were found at each locus in wheat. Both wheat Xwye838 variants, but only one of the Gss haplotypes seen in wheat, were found among the Ae. tauschii lines. The other wheat Gss haplotype was not found in either Ae. tauschii or 70 lines of tetraploid Ae. cylindrica (genomes CCDD), which is known to hybridize with wheat. It is concluded that both T. aestivum and Ae. cylindrica originated recurrently, with at least two genetically distinct progenitors contributing to the formation of the D genome in both species.

Base Sequence↗

PubMed2004

Deletion polymorphism in wheat chromosome regions with contrasting recombination rates.

Polymorphism for deletions was investigated in 1027 lines of tetraploid and hexaploid wheat and 420 lines of wheat diploid ancestors. A total of 26 deletions originating during the evolution of polyploid wheat were discovered among 155 investigated loci. Wheat chromosomes were divided into a proximal, low-recombination interval containing 69 loci and a distal, high-recombination interval containing 86 loci. A total of 23 deletions involved loci in the distal, high-recombination interval and only 3 involved loci in the proximal, low-recombination interval. The rates of DNA loss differed by several orders of magnitude in the two intervals. The rate of diploidization of polyploid wheat by deletions was estimated and was shown to have proceeded faster in the distal, high-recombination interval than in the proximal, low-recombination interval.

Evolution, Molecular↗

PubMed2003

High-throughput fingerprinting of bacterial artificial chromosomes using the snapshot labeling kit and sizing of restriction fragments by capillary electrophoresis.

We have developed an automated, high-throughput fingerprinting technique for large genomic DNA fragments suitable for the construction of physical maps of large genomes. In the technique described here, BAC DNA is isolated in a 96-well plate format and simultaneously digested with four 6-bp-recognizing restriction endonucleases that generate 3' recessed ends and one 4-bp-recognizing restriction endonuclease that generates a blunt end. Each of the four recessed 3' ends is labeled with a different fluorescent dye, and restriction fragments are sized on a capillary DNA analyzer. The resulting fingerprints are edited with a fingerprint-editing computer program and contigs are assembled with the FPC computer program. The technique was evaluated by repeated fingerprinting of several BACs included as controls in plates during routine fingerprinting of a BAC library and by reconstruction of contigs of rice BAC clones with known positions on rice chromosome 10.

Chromosome Mapping↗