RNA purified from the feline calicivirus (FCV) F9 vaccine strain was used to prepare a cDNA library in the expression vector lambda gt11. The library was screened for expression of FCV antigen using a rabbit antiserum prepared against purified FCV. A 330 bp cDNA clone was identified and used as a probe to obtain a larger overlapping clone of 1369 bp. Comparative sequence analysis with the CFI a...

Applications that use Bacterial Artificial Chromosome (BAC) libraries often require paired-end sequences and knowledge of the physical location of each clone in plates. To facilitate obtaining this information in high-throughput, we generated pBACode vectors: a pool of BAC cloning vectors, each with a pair of random barcodes flanking its cloning site. In a pBACode BAC library, the BAC ends and ...

One of the fundamental techniques of molecular biology is the isolation of a rare clone from a complex mixture of DNA sequences contained within a library. The isolation of cDNA and genomic clones from highly complex libraries is often an early step in studies aimed at understanding basic biological processes as well as in applied biological research. In a typical genomic library with an averag...

A problem in many sequencing projects is the final closure of gaps left in the clone libraries, which serve as templates for sequencing, because of uncloned or unclonable genomic areas. By use of the Xylella fastidiosa genome as a test system, we present here an approach to generate, in a directed manner, sequence information from those gaps. We suggest using the complete clone library as a com...

A Cys endopeptidase (SH-EP) is involved in the degradation of storage globulin in cotyledons of germinating seeds of Vigna mungo (Mitsuhashi and Minamikawa, 1989). The structure and expression of the gene for SH-EP have been reported (Akasofu et al., 1990; Yamauchi et al., 1992). Here we isolated a full-length cDNA for a putative Cys protease from a rice seed cDNA expression library using antis...

A clone bearing the structural gene sacB, coding for the exoenzyme levansucrase, was isolated from a library of Bacillus subtilis DNA that was cloned in phage lambda charon 4A on the basis of the transforming activity of the chimeric DNA. This lambda clone also was found to contain the sacR and smo loci. Subcloning the sacB-sacR region in plasmid pBR325 resulted in a clone which directed levans...

The nuclear yeast two-hybrid (Y2H) system is the most widely used technology for detecting interactions between proteins. A common approach is to screen specific test proteins (baits) against large compilations of randomly cloned proteins (prey libraries). For eukaryotic organisms, libraries have traditionally been generated using messenger RNA (mRNA) extracted from various tissues and cells. H...

To allow the efficient construction of ordered clone libraries, we have been investigating the use of 'oligonucleotide fingerprinting' as an approach to identify overlapping clones, and ultimately restore the linear order of the clone set. To test the effectiveness of the procedure, we have constructed a cosmid library from the genome of the human DNA virus HSV-I and used hybridisation to multi...

To allow the efficient construction of ordered clone libraries, we have been investigating the use of 'oligonucleotide fingerprinting' as an approach to identify overlapping clones, and ultimately restore the linear order of the clone set. To test the effectiveness of the procedure, we have constructed a cosmid library from the genome of the human DNA virus HSV-1 and used hybridisation to multi...