Identification and Cloning of Differentially Expressed Genes by Long-Distance Differential Display

Changes in transcription profiles reflect strain contributions to defined cultures of Lactococcus lactis subsp. cremoris during milk fermentation

“…The effect of strain association was demonstrated by distinguishing whole transcriptome profiles obtained with pure cultures from those obtained with defined mixed cultures. FRAP-PCR is not handicapped by problems of irreproducibility inherent to arbitrary PCR (Jurecic et al 1998;Zhao et al 1995), and does not show technical bias due to competition during PCR. Previous false positive problems could have been caused principally by the use of acrylamide electrophoresis, as many different amplimers (differentially expressed and constitutive) comigrate in the same band on gel (Bauer et al 1994).…”

Applications of Differential-Display Reverse Transcription-PCR to Molecular Pathogenesis and Medical Mycology

“…The lengthening of primers increases product size, resulting in longer sequences that can demonstrate homology to sequences reported in the database (67). Longer primers also improve direct sequencing of DDRT-PCR products (27,67,79,88,148). Either the anchor or the arbitrary primer can be end labeled and used to sequence the PCR product.…”

“…If the product is to be characterized further, the restriction enzyme sites on both ends facilitate directional cloning (148). Laboratories report between 10 and 50% success rates with direct cloning (67,88).…”

“…A common application of DDRT-PCR is to identify genes that are regulated during differentiation or development, including hematopoiesis (17,46,57,67,68,123), neuronal cell differentiation (13,64,153) senescence (79), and adipocyte differentiation (32). Normally, cell populations at different stages of development are compared.…”

Messenger RNA differntial display strategies in birds and amphibians

“…Since the initial publication, more than 1500 reports have described its application for the identification of differentially expressed genes in a wide range of biological systems. Many improvements to the method have been proposed, in particular addressing issues such as reducing falsepositive clones [2 -4], enhancing long-range amplifications [5], and systematically surveying the repertoire of differentially expressed mRNAs [4,6]. It would be beyond the scope of this report to describe the DD method in detail, and I will rather schematize an integrated strategy used in our laboratory which facilitates its use as a robust molecular screen for the detection of differentially regulated mRNAs, and their characterization as expressed sequence tags (ESTs).…”