Abstract：
BACKGROUND:The majority of protein isoforms arise from alternative splicing of the encoding primary RNA transcripts. To understand the significance of single splicing events, reliable techniques are needed to determine their incidence. However, existing methods are labour-intensive, error-prone or of limited use. RESULTS:Here, we present an improved method to determine the relative incidence of transcripts that arise from alternative splicing at a single site. Splice variants were quantified within a single sample using one-step reverse transcription quantitative PCR. Amplification products obtained with variant specific primer pairs were compared to those obtained with primer pairs common to both variants. The identities of variant specific amplicons were simultaneously verified by melt curve analysis. Independent calculations of the relative incidence of each variant were performed. Since the relative incidences of variants have to add upto 100%, the method provides an internal control to monitor experimental errors and uniform reverse transcription. The reliability of the method was tested using mixtures of cDNA templates as well as RNA samples from different sources. CONCLUSION:The method described here, is easy to set up and does not need unrelated reference genes and time consuming, error-prone standard curves. It provides a reliable and precise technique to distinguish small differences of the relative incidence of two splice variants.
journal_name
BMC Mol Bioljournal_title
BMC molecular biologyauthors
Camacho Londoño J,Philipp SEdoi
10.1186/s12867-016-0060-1subject
Has Abstractpub_date
2016-03-15 00:00:00pages
8issn
1471-2199pii
10.1186/s12867-016-0060-1journal_volume
17pub_type
杂志文章abstract：BACKGROUND:Senescence marker protein-30 (SMP30), whose expression declines during aging in rat liver, has been proposed as an important aging marker. Besides apoptosis, SMP30 also protects cells against various other injuries by enhancement of membrane calcium-pump activity. The mechanism of this differential gene expr...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-9-43
更新日期：2008-04-29 00:00:00
abstract：BACKGROUND:The challenge of determining amplification efficiency has long been a predominant aspect of implementing real-time qPCR, playing a critical role in the accuracy and reliability that can be achieved. Based upon analysis of amplification profile position, standard curves are currently the gold standard for amp...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-9-96
更新日期：2008-10-30 00:00:00
abstract：BACKGROUND:Visceral leishmaniasis caused by members of the Leishmania donovani complex is often fatal in the absence of treatment. Research has been hampered by the lack of good laboratory models and tools for genetic manipulation. In this study, we have characterised a L. infantum line (JPCM5) that was isolated from a...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-7-42
更新日期：2006-11-13 00:00:00
abstract：BACKGROUND:Retinal ganglion cells (RGCs) are preferentially lost in glaucoma or optic neuritis. In the present study, we investigated the protective effect of mircoRNA 100 (miR-100) against oxidative stress induced apoptosis in RGC-5 cells. RESULTS:Rat RGC-5 cells were cultured in plates and H2O2 was added to induce o...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/s12867-014-0025-1
更新日期：2014-11-18 00:00:00
abstract：BACKGROUND:The rapid increase in whole genome fungal sequence information allows large scale functional analyses of target genes. Efficient transformation methods to obtain site-directed gene replacement, targeted over-expression by promoter replacement, in-frame epitope tagging or fusion of coding sequences with fluor...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-9-70
更新日期：2008-08-01 00:00:00
abstract：BACKGROUND:The Y chromosome of mammals is particularly prone to accumulate genes related to male fertility. However, the high rate of molecular evolution on this chromosome predicts reduced power to the across-species comparative approach in identifying male-specific genes that are essential for sperm production in hum...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-11-33
更新日期：2010-05-07 00:00:00
abstract：BACKGROUND:Selecting stably expressed reference genes is essential for proper reverse transcription quantitative polymerase chain reaction gene expression analysis. However, this choice is not always straightforward. In the case of differentiating human embryonic stem (hES) cells, differentiation itself introduces chan...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-14-21
更新日期：2013-09-12 00:00:00
abstract：BACKGROUND:We reported previously that the cardiac troponin I (cTnI) promoter drives cardiac-specific expression of reporter genes in cardiac muscle cells and in transgenic mice, and that disruption of GATA elements inactivates the cTnI promoter in cultured cardiomyocytes. We have now examined the role of cTnI promoter...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-8-78
更新日期：2007-09-17 00:00:00
abstract：BACKGROUND:Cryptochromes (CRYs) are a class of flavoprotein blue-light signaling receptors found in plants and animals, and they control plant development and the entrainment of circadian rhythms. They also act as integral parts of the central circadian oscillator in humans and other animals. In mammals, the CLOCK-BMAL...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-11-69
更新日期：2010-09-14 00:00:00
abstract：BACKGROUND:Prevalence of fibroproliferative diseases, including chronic kidney disease is rapidly increasing and has become a major public health problem worldwide. Fibroproliferative diseases are characterized by increased expression of α smooth muscle actin (α-SMA) that belongs to the family of the six conserved acti...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/s12867-017-0089-9
更新日期：2017-04-27 00:00:00
abstract：BACKGROUND:Although staphylokianse (SAK) is among the most promising blood dissolving agents, it is far from ideal. It is interesting to hypothesize that the clot lysis efficacy of SAK can be enhanced with direct active platelet binding ability, and at the same time the rethrombosis complication after successful recana...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-8-88
更新日期：2007-10-07 00:00:00
abstract：BACKGROUND:Gene conversion depends upon the same factors that carry out more general process of homologous recombination, including homologous gene targeting and recombinational repair. Among these are the RAD51 paralogs, conserved factors related to the key recombination factor, RAD51. In chicken and other fowl, gene ...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-10-98
更新日期：2009-10-28 00:00:00
abstract：BACKGROUND:Cdc23/Mcm10 is required for the initiation and elongation steps of DNA replication but its biochemical function is unclear. Here, we probe its function using a novel approach in fission yeast, involving Cdc23 cleavage by the TEV protease. RESULTS:Insertion of a TEV protease cleavage site into Cdc23 allows i...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-6-13
更新日期：2005-06-07 00:00:00
abstract：BACKGROUND:The enzyme glucose-6-phosphatase catalyzes the dephosphorylation of glucose-6-phosphatase to glucose, the final step in the gluconeogenic and glycogenolytic pathways. Expression of the glucose-6-phosphatase gene is induced by glucocorticoids and elevated levels of intracellular cAMP. The effect of cAMP in re...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-6-2
更新日期：2005-01-19 00:00:00
abstract：BACKGROUND:Single-stranded DNA-binding proteins are essential cellular components required for the protection, metabolism and processing of single-stranded DNA. Human single-stranded DNA-binding protein 1 (hSSB1) is one such protein, with described roles in genome stability maintenance and transcriptional regulation. A...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/s12867-016-0077-5
更新日期：2016-12-09 00:00:00
abstract：BACKGROUND:Nucleosomes are nucleoproteic complexes, formed of eight histone molecules and DNA, and they are responsible for the compaction of the eukaryotic genome. Their presence on DNA influences many cellular processes, such as transcription, DNA replication, and DNA repair. The evolutionarily conserved histone vari...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-10-18
更新日期：2009-03-04 00:00:00
abstract：BACKGROUND:Among the three functions of DNA, mammalian replication and transcription can be subject to epigenetic imprinting specified by the parental origin of chromosomes, and although there is suggestive indication that this is also true for meiotic recombination, no definitive evidence has yet been reported. RESUL...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-10-43
更新日期：2009-05-13 00:00:00
abstract：BACKGROUND:Small proline rich protein (SPRR) 2A is one of 14 SPRR genes that encodes for a skin cross-linking protein, which confers structural integrity to the cornified keratinocyte cell envelope. New evidence, however, shows that SPRR2A is also a critical stress and wound repair modulator: it enables a variety of ba...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-13-20
更新日期：2012-06-25 00:00:00
abstract：BACKGROUND:ALL1-fused from chromosome 1q (AF1q), originally considered as an oncogenic factor, has been implicated in neuronal development; however, its upstream regulatory mechanisms in neural system remained elusive. RESULTS:Our study showed that REST (RE1 silencing transcription factor), a key transcription factor ...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/s12867-015-0043-7
更新日期：2015-09-05 00:00:00
abstract：BACKGROUND:In spite of large intergenic spaces in plant and animal genomes, 7% to 30% of genes in the genomes encode overlapping cis-natural antisense transcripts (cis-NATs). The widespread occurrence of cis-NATs suggests an evolutionary advantage for this type of genomic arrangement. Experimental evidence for the regu...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-9-6
更新日期：2008-01-14 00:00:00
abstract：BACKGROUND:Poly(ADP-ribose) polymerase-1 (PARP-1) is a nuclear enzyme that plays critical functions in many biological processes, including DNA repair and gene transcription. The main function of PARP-1 is to catalyze the transfer of ADP-ribose units from nicotinamide adenine dinucleotide (NAD+) to a large array of acc...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-8-96
更新日期：2007-10-25 00:00:00
abstract：BACKGROUND:Core promoters are cis-regulatory modules to which bind the basal transcriptional machinery and which participate in the regulation of transcription initiation. Although core promoters have not been extensively investigated through functional assays in a chromosomal context, the available data suggested that...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-12-32
更新日期：2011-08-01 00:00:00
abstract：BACKGROUND:Trichomonosis, caused by Trichomonas vaginalis, is the number one, nonviral sexually transmitted infection that has adverse consequences for the health of women and children. The interaction of T. vaginalis with vaginal epithelial cells (VECs), a step preparatory to infection, is mediated in part by the prom...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-6-5
更新日期：2005-03-04 00:00:00
abstract：BACKGROUND:Krüppel-like factors (KLFs) are a group of master regulators of gene expression conserved from flies to human. However, scant information is available on either the mechanisms or functional impact of the coupling of KLF proteins to chromatin remodeling machines, a deterministic step in transcriptional regula...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-15-10
更新日期：2014-05-25 00:00:00
abstract：BACKGROUND:cDNA-RDA is one of the subtractive cloning techniques used to isolate differentially expressed genes between two complex cDNA populations. In the present study we present a modification of the protocol described by Hubank and Schatz. RESULTS:In the post-hybridization mix, the 5'-ends of homoduplexes of inte...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-3-6
更新日期：2002-04-24 00:00:00
abstract：BACKGROUND:Physical exercise can improve brain function by altering brain gene expression. The expression mechanisms underlying the brain's response to exercise still remain unknown. miRNAs as vital regulators of gene expression may be involved in regulation of brain genes in response to exercise. However, as yet, very...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/s12867-019-0120-4
更新日期：2019-01-15 00:00:00
abstract：BACKGROUND:The selection of stable and suitable reference genes for real-time quantitative PCR (RT-qPCR) is a crucial prerequisite for reliable gene expression analysis under different experimental conditions. The present study aimed to identify reference genes as internal controls for gene expression studies by RT-qPC...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-13-22
更新日期：2012-06-29 00:00:00
abstract：BACKGROUND:Double-strand breakage of chromosomal DNA is obviously a serious threat to cells because various activities of the chromosome depend on its integrity. However, recent experiments suggest that such breakage may occur frequently during "normal" growth in various organisms - from bacteria through vertebrates, p...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-4-5
更新日期：2003-04-28 00:00:00
abstract：BACKGROUND:Naturally occurring tRNAs contain numerous modified nucleosides. They are formed by enzymatic modification of the primary transcripts during the complex RNA maturation process. In model organisms Escherichia coli and Saccharomyces cerevisiae most enzymes involved in this process have been identified. Interes...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-7-23
更新日期：2006-07-18 00:00:00
abstract：BACKGROUND:Conserved two-component system ColRS of Pseudomonas genus has been implicated in several unrelated phenotypes. For instance, deficiency of P. putida ColRS system results in lowered phenol tolerance, hindrance of transposition of Tn4652 and lysis of a subpopulation of glucose-grown bacteria. In order to disco...
journal_title：BMC molecular biology
pub_type： 杂志文章
doi：10.1186/1471-2199-10-46
更新日期：2009-05-16 00:00:00
