PubMed Health⌕ Search

SEARCH · PubMed Health

Results for “Genomic Library”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources
PubMed1991

A DNA transformation-competent Arabidopsis genomic library in Agrobacterium.

We have constructed a nuclear genomic library from the cruciferous plant Arabidopsis thaliana ecotype Columbia in a cosmid vector, pLZO3, and a host organism, Agrobacterium tumefaciens AGL1, which can directly DNA-transform the parent organism, Arabidopsis. The broad host range cosmid pLZO3 carries a gentamicin acetyltransferase gene as bacterial selective marker and tandem, chimeric neomycin and streptomycin phosphotransferase genes as plant selective markers. Agrobacterium AGL1 carries the hypervirulent, attenuated tumor-inducing plasmid pTiBo542 from which T-region DNA sequences have been precisely deleted, allowing optimal DNA transformation of many dicotyledonous plants. Agrobacterium AGL1 also carries an insertion mutation in its recA general recombination gene, which stabilizes the recombinant plasmids. The Arabidopsis genomic library consists of some 21,600 clones gridded onto 96-well microtiter dishes and, if random, carries at least three genomic equivalents. When probed for the presence of several Arabidopsis low copy-number genes, the genomic library seems representative. As with the unicellular organisms Escherichia coli and Saccharomyces cerevisiae, this DNA transformation competent genomic library should expedite gene isolation, by gene rescue, in multicellular organisms like Arabidopsis.

Agrobacterium tumefaciens↗
PubMed1992

Derivation of a mathematical expression useful for the construction of complete genomic libraries.

We present and derive a formula that is useful for the design and evaluation of gene cloning experiments in which a complete gene library of the entire genome of an organism is desired. The formula n = ln(1-phi f)/ln(1-f) (in which n is the number of recombinant clones required to ensure a probability, phi, of obtaining at least one of each of all possible gene sequences, and f is the fraction of the genome contained in an average-sized DNA fragment) applies to construction of libraries, in which at least one copy of all the genetic information of a genome is required. The use of this formula for quantitative evaluation of genomic libraries should give greater assurance that a given library would be complete.

Cloning, Molecular↗
PubMed1992

[The construction of genomic library of Alcaligenes faecalis and cloning of nifH gene sequence homologous to Klebsiella pneumoniae].

A genomic library of Alcaligenes faecalis A-15Hl which possesses rather high nitrogenase activity has been constructed. The total DNA of A. faecalis A-15 Hl was partially digested with Sau3AI. 13-20 kb of fragments recovered from agarose gel were cloned in bacteriophage EM-BL4 vector. A total number of 1.2 x 10(6) of recombinants was obtained. It is much beyond the desired capacity of a library. By using nifH gene of K. pneumoniae from plasmid pGB1 as probe, we have successfully screened the clone containing its homologous sequence. The recombinant bacteriophage DNA was digested with EcoRI. A 3.5 kb of hybridizing band appeared after southern blotting and then was cloned in pUC19 DNA. The result of southern blotting indicated that the recombinant plasmid clone contained nifH gene sequence of A. faecalis. This clone was named as pAFH.

Alcaligenes↗
PubMed1992

A mouse genomic library in the bacteriophage P1 cloning system: organization and characterization.

Using the bacteriophage P1 cloning system, we have constructed a two to three times coverage, high-molecular-weight (HMW) genomic library from mouse C127 fibroblast cells. The library consists of about 127,500 clones with an average insert size of about 70 kb that are organized into 300 primary pools containing approximately 425 clones per pool. For screening purposes the primary pools are combined into secondary pools (approximately 4250 clones each) and tertiary pools (approximately 21,250 clones each). Screening is performed by the polymerase chain reaction (PCR) with DNA isolated from the secondary and tertiary pools. We have screened the library for 13 different mouse sequences and have detected 11. Clones generated from two of the eleven positive screens were isolated from the library (those containing the c-fos and G alpha i2 genes) and were further characterized. Direct double-stranded sequencing of DNA from P1 clones with primers bordering the insert provided sequence information from each end of the cloned DNA.

Animals↗
PubMed1992

Versatile shuttle vectors and genomic libraries for use with Schizosaccharomyces pombe.

We have constructed a variety of pUC-based vectors designed for maintenance in Schizosaccharomyces pombe. These can be used for both gene bank construction and subcloning. Plasmids pUR18 and pUR19 are modifications of pUC vectors containing the Sc. pombe ars1 and ura4 sequences and retaining the lacZ XGal blue-white selection system for screening for DNA inserts. These vectors have been used to construct representative Sc. pombe and Saccharomyces cerevisiae genomic libraries. To assist in the creation of gene deletions, we have constructed another two plasmids. Combined with the technique of partially filling-in 5' overhangs created with restriction enzymes, these plasmids simplify the replacement of all or part of an open reading frame by a functional ura4 gene. Furthermore, such constructs can be excised with SfiI as a linear fragment for use in Sc. pombe transformations. When integrated into the Sc. pombe genome, the site of integration can be easily mapped by pulsed-field gel electrophoresis using the presence of a novel NotI site.

Base Sequence↗
PubMed1992

The construction of Streptomyces cyaneus genomic libraries in Escherichia coli is dependent upon the use of Mcr-deficient strains.

Streptomyces cyaneus genomic DNA ligated into either lambda phage or plasmid vectors was very inefficiently cloned into standard Escherichia coli host strains. However, the same material could be efficiently cloned using Mcr-deficient E. coli strains. These results suggest that the S. cyaneus genome contains 5-methylcytosine residues, some of which occur within the recognition sequences of the E. coli Mcr restriction system.

Cloning, Molecular↗
PubMed1992

Isolation of chromosome 21-specific yeast artificial chromosomes from a total human genome library.

A new approach for the isolation of chromosome-specific subsets from a human genomic yeast artificial chromosome (YAC) library is described. It is based on the hybridization with an Alu polymerase chain reaction (PCR) probe. We screened a 1.5 genome equivalent YAC library of megabase insert size with Alu PCR products amplified from hybrid cell lines containing human chromosome 21, and identified a subset of 63 clones representative of this chromosome. The majority of clones were assigned to chromosome 21 by the presence of specific STSs and in situ hybridization. Twenty-nine of 36 STSs that we tested were detected in the subset, and a contig spanning 20 centimorgans in the genetic map and containing 8 STSs in 4 YACs was identified. The proposed approach can greatly speed efforts to construct physical maps of the human genome.

Base Sequence↗
PubMed1992

Dominant genetics using a yeast genomic library under the control of a strong inducible promoter.

In Saccharomyces cerevisiae, numerous genes have been identified by selection from high-copy-number libraries based on "multicopy suppression" or other phenotypic consequences of overexpression. Although fruitful, this approach suffers from two major drawbacks. First, high copy number alone may not permit high-level expression of tightly regulated genes. Conversely, other genes expressed in proportion to dosage cannot be identified if their products are toxic at elevated levels. This work reports construction of a genomic DNA expression library for S. cerevisiae that circumvents both limitations by fusing randomly sheared genomic DNA to the strong, inducible yeast GAL1 promoter, which can be regulated by carbon source. The library obtained contains 5 x 10(7) independent recombinants, representing a breakpoint at every base in the yeast genome. This library was used to examine aberrant gene expression in S. cerevisiae. A screen for dominant activators of yeast mating response identified eight genes that activate the pathway in the absence of exogenous mating pheromone, including one previously unidentified gene. One activator was a truncated STE11 gene lacking approximately 1000 base pairs of amino-terminal coding sequence. In two different clones, the same GAL1 promoter-proximal ATG is in-frame with the coding sequence of STE11, suggesting that internal initiation of translation there results in production of a biologically active, truncated STE11 protein. Thus this library allows isolation based on dominant phenotypes of genes that might have been difficult or impossible to isolate from high-copy-number libraries.

Base Sequence↗
PubMed1992

[The construction of rich genomic library of bovine Y chromosome].

According to the fundamentals of DNA renatured kinetics, we made a study in which genomic DNA from male cattle was completely digested with Sau 3A, while female DNA was sheared to an average size of 500 bp fragments. The Y derived DNA sequence rich library of bovine has been contracted with pUC19 plasmid as vector and JM109 as host bacteria by reassociation of male and female DNA. Male and female DNA probes, were used. The result of colony hybridization showed that out of 1000 recombinants analysed, 6 were found to be stronger signal with male probe. Analysis of Pvu' I digested DNA fragment revealed that most of the inserts were in the 90-300 bp.

Animals↗
PubMed1992

Construction of small-insert genomic DNA libraries highly enriched for microsatellite repeat sequences.

We describe an efficient method for the construction of small-insert genomic libraries enriched for highly polymorphic, simple sequence repeats. With this approach, libraries in which 40-50% of the members contain (CA)n repeats are produced, representing an approximately 50-fold enrichment over conventional small-insert genomic DNA libraries. Briefly, a genomic library with an average insert size of less than 500 base pairs was constructed in a phagemid vector. Amplification of this library in a dut ung strain of Escherichia coli allowed the recovery of the library as closed circular single-stranded DNA with uracil frequently incorporated in place of thymine. This DNA was used as a template for second-strand DNA synthesis, primed with (CA)n or (TG)n oligonucleotides, at elevated temperatures by a thermostable DNA polymerase. Transformation of this mixture into wild-type E. coli strains resulted in the recovery of primer-extended products as a consequence of the strong genetic selection against single-stranded uracil-containing DNA molecules. In this manner, a library highly enriched for the targeted microsatellite-containing clones was recovered. This approach is widely applicable and can be used to generate marker-selected libraries bearing any simple sequence repeat from cDNAs, whole genomes, single chromosomes, or more restricted chromosomal regions of interest.

Animals↗
PubMed1992

Construction of a genomic DNA library of the Toxoplasma gondii ZS2 strain, screening of specific clones, and DNA diagnosis of toxoplasmosis.

We have constructed a genomic DNA library of the Toxoplasma gondii ZS2 strain and isolated a specific cloned DNA sequence from this organism. The restriction map of this cloned 1.1-kb DNA fragment was analyzed. Southern and dot-blot analyses showed that the 32P-labeled DNA fragment hybridized to parasite DNA, to DNAs from peripheral blood leukocytes and the thymus of baby pigs that were artificially infected with T. gondii, and to DNAs of T. gondii-positive anencephalic and hydrocephalic fetuses. It did not hybridize with DNA from controls, (i.e., normal human and baby pig peripheral blood leukocytes, spleen of normal mice, Plasmodium falciparum, Pneumocystis carinii, and pBR322). As few as 100 T. gondii parasites or 500 pg of purified DNA from T. gondii can be detected by dot-blot hybridization. This probe method was specific and sensitive, and has been used successfully in detecting various clinical cases of toxoplasmosis with T. gondii.

Animals↗
PubMed1992

Cloning and characterization of the human thyroid hormone receptor beta 1 gene promoter.

The promoter region of the human (h) thyroid hormone receptor (TR) beta 1 gene was isolated from a human placenta genomic library. Primer extension and S1 nuclease mapping confirmed a single transcriptional start site. DNA sequence analysis of the 5' upstream region revealed the existence of a putative thyroid response element (TRE) which is highly homologous to TREs found in several thyroid hormone responsive genes. Binding of hTR protein to the promoter region of the hTR beta 1 gene was confirmed by gel mobility shift assay. A transient transfection study demonstrated that hTR activated the expression of a reporter gene containing the promoter sequence of the hTR beta 1 gene in a hormone dependent manner. The TRE in the hTR beta 1 gene promoter may be involved in the autoregulation of hTR beta 1 gene expression.

Base Sequence↗
PubMed1992

Characterization of swine short interspersed repetitive sequences.

Swine genomic DNA segments containing repetitive sequences were isolated from a porcine genomic library using genomic DNA as a probe. Three fragments containing the repetitive sequences from two of the primary phage clones were subcloned for sequence analysis, which revealed six new PRE-1 repetitive families other than those reported earlier by Singer et al. (Nucleic Acids Research 15, 2780, 1987). The frequency of the repetitive sequences in the swine genome was estimated at 2 x 10(6) per diploid genome. Sequence analysis revealed similarities between these repetitive sequences and that of arginine-tRNA gene.

Animals↗
PubMed1992

Development of a polymerase chain reaction-based diagnosis of Trichomonas vaginalis.

We developed a polymerase chain reaction (PCR)-based test for detecting the protozoan parasite Trichomonas vaginalis. Genomic libraries were constructed from two independent clinical isolates of T. vaginalis. From these libraries, 12 genomic clones were purified, sequenced, and then screened for uniqueness by computer-assisted sequence comparisons. PCR reactions were performed to evaluate eight PCR-primer pairs, including a primer pair that targeted the T. vaginalis ferredoxin gene. All eight primer pairs yielded PCR products of the expected sizes. However, six of the primer pairs amplified their respective target sequences in limited numbers of clinical T. vaginalis isolates, suggesting the presence of significant genomic variability among isolates. An exception was a primer pair, termed TVA5-TVA6, that amplified a 102-bp genomic sequence, termed A6p, in all of 24 clinical isolates. The A6p sequence was not detected by PCR in human DNA or in a wide variety of flagellates, ciliates, or bacteria tested. The A6p sequence appears highly selective for a broad range of T. vaginalis isolates and holds promise for PCR-based diagnosis of the parasite.

Animals↗
PubMed1992

Differential molecular mechanism of the estrogen action that regulates lactoferrin gene in human and mouse.

The 5'-flanking region of the human lactoferrin gene was isolated from a human placental genomic library. This genomic clone contains a 16-kilobase pair (kbp) insert and produces seven fragments when digested with the SacI restriction enzyme. We sequenced one of the fragments that comprises 1294 bp of the 5'-flanking sequence, 79 bp of the first exon, and 690 bp of the first intron. A major transcription start site was mapped by primer extension. The region immediately upstream from the transcription initiation site following the first exon is abundant in G and C nucleotides. In the promoter and 5'-flanking region within a 300-bp stretch (-465 to -165) of the DNA, we found a noncanonical TATA box (ATAAA), CAAT-like sequence (CAAC) and sequences homologous to the consensus SP1 binding site, Pu.1/Sp.1 binding element (PU box), two half-palindromic estrogen response elements (EREs; GGTCA), an imperfect ERE (GGTCAAGGCGATC), and a sequence resembling the chicken ovalbumin upstream promoter transcription factor (COUP-TF) binding site (GTCTCACAGGTCA). The COUP-TF binding site and the imperfect ERE shared five nucleotides (GGTCA). With the exception of the two half-palindromic EREs, the elements with very well matched sequences were also found in the corresponding positions in the mouse lactoferrin gene. The synthetic oligonucleotide, including the 26 bp of COUP/ERE sequence, was cloned before the SV40 promoter in a chloramphenicol acetyltransferase reporter construct. These chimeric plasmids were transiently transfected into human endometrium carcinoma RL95-2 cells to assess hormone responsiveness. We found that the COUP/ERE element acted as an enhancer in response to estrogen stimulation. In vitro DNase I footprinting analysis showed binding of the estrogen receptor on the imperfect ERE. In contrast to the mouse lactoferrin COUP/ERE element, COUP-TF does not interact with this element, as demonstrated by band shift assay and site-directed mutagenesis. Therefore, the molecular mechanisms of the estrogen action that govern the lactoferrin gene expression differ between mouse and human.

Amino Acid Sequence↗
PubMed1992

The CEPH YAC library.

Because of their large size, YACs provide is a powerful tool for physical mapping studies of complex genomes. As it will be advantageous to have genomic libraries of clones with large inserts for analyzing megabase sized regions of the human genome, we have investigated a number of parameters in order to increase the insert size of the YACs. We constructed a genomic library currently containing more than 85,000 YAC clones. Mean sizes of YACs produced at several stages of construction of the library range from 430 kb to 1,200 kb, representing 13 haploid equivalents of the human genome. This library was organized in order to allow rapid screen of YACs for large scale physical mapping of the human genome.

Chromosome Walking↗
PubMed2025

RabbitSketch: a high-performance sketching library for genome analysis.

SUMMARY: We present RabbitSketch, a highly optimized library of sketching algorithms such as MinHash, OrderMinHash, and HyperLogLog that can exploit the power of modern multi-core CPUs. It provides significant speedups compared to existing implementations, ranging from 2.30× to 49.55×, as well as flexible and easy-to-use interfaces for both Python and C++. As a result, the similarity analysis of 455GB genomic data can be completed in only 5 minutes using RabbitSketch with merely 20 lines of Python code. As a case study, we enhanced RabbitTClust by integrating RabbitSketch's Kssd algorithm, resulting in a 1.54× speedup with no loss in accuracy. AVAILABILITY AND IMPLEMENTATION: RabbitSketch is available at https://github.com/RabbitBio/RabbitSketch with an archived version at Zenodo: https://doi.org/10.5281/zenodo.14903962. Detailed API documentation is available at https://rabbitsketch.readthedocs.io/en/latest.

Software↗