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At least 793 records · Page 44Linked to original sources
PubMed2006

Regulation of AUXIN RESPONSE FACTOR3 by TAS3 ta-siRNA affects developmental timing and patterning in Arabidopsis.

MicroRNAs (miRNAs) and trans-acting siRNAs (ta-siRNAs) in plants form through distinct pathways, although they function as negative regulators of mRNA targets by similar mechanisms . Three ta-siRNA gene families (TAS1, TAS2, and TAS3) are known in Arabidopsis thaliana. Biogenesis of TAS3 ta-siRNAs, which target mRNAs encoding several AUXIN RESPONSE FACTORs (including ARF3/ETTIN and ARF4 ) involves miR390-guided processing of primary transcripts, conversion of a precursor to dsRNA through RNA-DEPENDENT RNA POLYMERASE6 (RDR6) activity, and sequential DICER-LIKE4 (DCL4)-mediated cleavage events. We show that the juvenile-to-adult phase transition is normally suppressed by TAS3 ta-siRNAs, in an ARGONAUTE7-dependent manner, through negative regulation of ARF3 mRNA. Expression of a nontargeted ARF3 mutant (ARF3mut) in a wild-type background reproduced the phase-change phenotypes detected in rdr6-15 and dcl4-2 mutants, which lose all ta-siRNAs. Expression of either ARF3 or ARF3mut in rdr6-15 plants, in which both endogenous and transgenic copies of ARF3 were derepressed, resulted in further acceleration of phase change and severe morphological and patterning defects of leaves and floral organs. In light of the functions of ARF3 and ARF4 in organ asymmetry, these data reveal multiple roles for TAS3 ta-siRNA-mediated regulation of ARF genes in developmental timing and patterning.

Arabidopsis↗
PubMed2006

"Self" and "nonself" manipulation of interferon defense during persistent infection: bovine viral diarrhea virus resists alpha/beta interferon without blocking antiviral activity against unrelated viruses replicating in its host cells.

Bovine viral diarrhea virus (BVDV), together with Classical swine fever virus (CSFV) and Border disease virus (BDV) of sheep, belongs to the genus Pestivirus of the Flaviviridae. BVDV is either cytopathic (cp) or noncytopathic (ncp), as defined by its effect on cultured cells. Infection of pregnant animals with the ncp biotype may lead to the birth of persistently infected calves that are immunotolerant to the infecting viral strain. In addition to evading the adaptive immune system, BVDV evades key mechanisms of innate immunity. Previously, we showed that ncp BVDV inhibits the induction of apoptosis and alpha/beta interferon (IFN-alpha/beta) synthesis by double-stranded RNA (dsRNA). Here, we report that (i) both ncp and cp BVDV block the induction by dsRNA of the Mx protein (which can also be induced in the absence of IFN signaling); (ii) neither biotype blocks the activity of IFN; and (iii) once infection is established, BVDV is largely resistant to the activity of IFN-alpha/beta but (iv) does not interfere with the establishment of an antiviral state induced by IFN-alpha/beta against unrelated viruses. The results of our study suggest that, in persistent infection, BVDV is able to evade a central element of innate immunity directed against itself without generally compromising its activity against unrelated viruses ("nonself") that may replicate in cells infected with ncp BVDV. This highly selective "self" and "nonself" model of evasion of the interferon defense system may be a key element in the success of persistent infection in addition to immunotolerance initiated by the early time point of fetal infection.

Animals↗
PubMed2006

The PDZ-GEF dizzy regulates cell shape of migrating macrophages via Rap1 and integrins in the Drosophila embryo.

In Drosophila embryos, macrophages originate from the cephalic mesoderm and perform a complex migration throughout the entire embryo. The molecular mechanisms regulating this cell migration remain largely unknown. We identified the Drosophila PDZ G-nucleotide exchange factor (PDZ-GEF) Dizzy as a component essential for normal macrophage migration. In mutants lacking Dizzy, macrophages have smaller cellular protrusions, and their migration is slowed down significantly. This phenotype appears to be cell-autonomous, as it is also observed in embryos with a dsRNA-induced reduction of dizzy function in macrophages. In a complementary fashion, macrophages overexpressing Dizzy are vastly extended and form very long protrusions. These cell shape changes depend on the function of the small GTPase Rap1: in rap1 mutants, Dizzy is unable to induce the large protrusions. Furthermore, forced expression of a dominant-active form of Rap1, but not of the wild-type form, induces similar cell shape changes as Dizzy does overexpression. These findings suggest that Dizzy acts through Rap1. We propose that integrin-dependent adhesion is a Rap1-mediated target of Dizzy activity: in integrin mutants, neither Dizzy nor Rap1 can induce cell shape changes in macrophages. These data provide the first link between a PDZ-GEF, the corresponding small GTPase and integrin-dependent cell adhesion during cell migration in embryonic development.

Animals↗
PubMed2001

RNA-dependent protein kinase PKR is required for activation of NF-kappa B by IFN-gamma in a STAT1-independent pathway.

The IFN-inducible dsRNA-activated protein kinase PKR regulates protein synthesis through phosphorylation of eukaryotic initiation factor-2alpha. It also acts as a signal transducer for transcription factors NF-kappaB, IFN regulatory factor-1, and activating transcription factor-2. IFN-gamma, a pleiotropic cytokine, elicits gene expression by activating the Janus kinase-STAT signaling pathway. IFN-gamma can synergize with TNF-alpha to activate NF-kappaB in a number of cell lines. Here we show that IFN-gamma alone can activate NF-kappaB, by a Janus kinase-1-mediated, but Stat1-independent, mechanism. NF-kappaB activation by IFN-gamma is associated with degradation of IkappaB beta. The IFN-gamma response can be blocked by 2',5'-oligoadenylate-linked antisense chimeras against PKR mRNA. There was no activation of NF-kappaB by IFN in PKR-null cells, indicating that PKR is required for IFN-gamma signaling to NF-kappaB.

3T3 Cells↗
PubMed2005

Analysis of genes induced by Sendai virus infection of mutant cell lines reveals essential roles of interferon regulatory factor 3, NF-kappaB, and interferon but not toll-like receptor 3.

Sendai virus (SeV) infection causes the transcriptional induction of many cellular genes that are also induced by interferon (IFN) or double-stranded RNA (dsRNA). We took advantage of various mutant cell lines to investigate the putative roles of the components of the IFN and dsRNA signaling pathways in the induction of those genes by SeV. Profiling the patterns of gene expression in SeV-infected cells demonstrated that Toll-like receptor 3, although essential for gene induction by dsRNA, was dispensable for gene induction by SeV. In contrast, Jak1, which mediates IFN signaling, was required for the induction of a small subset of genes by SeV. NF-kappaB and interferon regulatory factor 3 (IRF-3), the two major transcription factors activated by virus infection, were essential for the induction of two sets of genes by SeV. As expected, some of the IRF-3-dependent genes, such as ISG56, were more strongly induced by SeV in IRF-3-overexpressing cells. Surprisingly, in those cells, a number of NF-kappaB-dependent genes, such as the A20 gene, were induced poorly. Using a series of cell lines expressing increasing levels of IRF-3, we demonstrated that the degree of induction of A20 mRNA, upon SeV infection, was inversely proportional to the cellular level of IRF-3, whereas that of ISG56 mRNA was directly proportional. Thus, IRF-3 can suppress the expression of NF-kappaB-dependent genes in SeV-infected cells.

Cell Line↗
PubMed2005

Serum-dependent expression of promyelocytic leukemia protein suppresses propagation of influenza virus.

The rate of propagation of influenza virus in human adenocarcinoma Caco-2 cells was found to negatively correlate with the concentration of fetal bovine serum (FBS) in the culture medium. Virus replicated more rapidly at lower FBS concentrations (0 or 2%) than at higher concentrations (10 or 20%) during an early stage of infection. Basal and interferon (IFN)-induced levels of typical IFN-inducible anti-viral proteins, such as 2',5'-oligoadenylate synthetase, dsRNA-activated protein kinase and MxA, were unaffected by variation in FBS concentrations. But promyelocytic leukemia protein (PML) was expressed in a serum-dependent manner. In particular, the 65 to 70 kDa isoform of PML was markedly upregulated following the addition of serum. In contrast, other isoforms were induced by IFN treatment, and weakly induced by FBS concentrations. Immunofluorescence microscopy indicated that PML was mainly formed nuclear bodies in Caco-2 cells at various FBS concentrations, and the levels of the PML-nuclear bodies were upregulated by FBS. Overexpression of PML isoform consisting of 560 or 633 amino acid residues by transfection of expression plasmid results in significantly delayed viral replication rate in Caco-2 cells. On the other hand, downregulation of PML expression by RNAi enhanced viral replication. These results indicate that PML isoforms which are expressed in a serum-dependent manner suppress the propagation of influenza virus at an early stage of infection.

2',5'-Oligoadenylate Synthetase↗
PubMed2002

Interferon regulatory factor-1, interferon-beta, and reovirus-induced myocarditis.

Viral myocarditis is an important human disease, and reovirus-induced myocarditis in mice provides an excellent model to study direct viral damage to the heart. Previously, we showed that reovirus induction of and sensitivity to interferon-beta (IFN-beta) is an important determinant of viral pathogenicity in the heart and that the transcription factor interferon regulatory factor-3 (IRF-3) is required for reovirus induction of IFN-beta in primary cardiac myocyte cultures. Given several lines of evidence suggesting a possible distinctive environment for IRFs in the heart, we have now focused on IRF-1. Previous studies demonstrated that viruses, double-stranded-RNA (dsRNA), and IFN-alpha/beta can each induce IRF-1 and that IRF-1 plays a role in dsRNA, but perhaps not viral, induction of IFN-alpha/beta. Importantly, none of these studies used a virus with a dsRNA genome (such as reovirus), none of them used a highly differentiated nonlymphoid cell type, and none of them addressed whether viral induction of IRF-1 is direct or is mediated through viral induction of IFN-beta. Indeed, as recently as this year it has been assumed that viral induction of IRF-1 is direct. Here, we found that reovirus induced IRF-1 in primary cardiac myocyte cultures, but that IRF-1 was not required for reovirus induction of IFN-beta. Surprisingly, we found that reovirus failed to induce IRF-1 in the absence of the IFN-alpha/beta response. This provides the first evidence that viruses may not induce IRF-1 directly. Finally, nonmyocarditic reovirus strains induced more cardiac lesions in mice deficient for IRF-1 than they did in wildtype mice, directly demonstrating a protective role for IRF-1. Together, the results indicate that while IRF-1 is downstream of the IFN-beta response, it plays an important protective role against viral myocarditis.

Animals↗
PubMed1994

Proteins that interact with PKR.

The in vitro activities of recombinant gene products of the vaccinia virus E3L and K3L genes have been compared. These proteins are both potent inhibitors of the dsRNA activated protein kinase (PKR) as assayed in cell-free translation systems or with purified PKR. The two gene products function at similar molar concentrations. Both proteins are expressed early in vaccinia virus infection suggesting that vaccinia virus maintains redundant mechanisms for the down regulation of PKR. The K3L gene product can be shown to be associated with PKR in vaccinia virus infected cells. The activities of the vaccinia virus PKR inhibitors are compared with other viral protein inhibitors of PKR. A variety of cellular proteins have also been identified by their ability to inhibit PKR activity or to prevent PKR activation. These cellular PKR interacting proteins have been uncovered from the studies of viral strategies to prevent PKR activation, as well as from studies looking at the effects of growth control, growth factors or oncogene expression on PKR activity. A picture emerges of PKR fulfilling a complex regulatory role in cell function with the regulation of its activity as part of a complex cascade interfacing with the signal transduction/cell cycle control machinery.

Animals↗
PubMed2002

An antiviral state induced in Chinook salmon embryo cells (CHSE-214) by transfection with the double-stranded RNA poly I:C.

Type I interferons (IFN alpha and beta) convert vertebrate cells into an antiviral state by inducing expression of proteins that inhibit virus replication. In humans and mice, Mx proteins constitute one family of interferon-induced antiviral proteins. Mx genes have recently been cloned from Atlantic salmon and rainbow trout. Moreover, double-stranded RNA (dsRNA) and type I IFN-like activity have been shown to induce Mx protein in salmonid cells. Chinook salmon embryo cells (CHSE-214 cells) have been suggested to have a defect in the IFN-system because the dsRNA polyinosinic polycytidylic acid (poly I:C) failed to induce an antiviral state in the cells. We have studied this phenomenon more closely in the present work. CHSE-214 cells were either transfected with poly I:C or incubated with poly I:C without transfection reagent. The cells were then studied for Mx protein expression and protection against infectious pancreatic necrosis virus (IPNV) infection. The results showed that cells transfected with poly I:C were protected from IPNV infection, whilst cells incubated with poly I:C were not protected. Cells transfected with the double-stranded DNA poly dI:dC were also not protected against IPNV. Mx protein was expressed in CHSE-214 cells upon transfection with poly I:C, but not after incubation with poly I:C alone. Stimulation of CHSE-214 cells with supernatants from cells transfected with poly I:C, induced protection against IPNV, indicating production of type I IFN-like activity. These results suggest that CHSE-214 cells in fact are able to produce type I IFN, but may have defects in the mechanisms mediating uptake of poly I:C or may degrade unprotected poly I:C.

Animals↗
PubMed2004

Transgenic RNAi-mediated reduction of MSY2 in mouse oocytes results in reduced fertility.

MSY2 is implicated in regulating the stability and translation of maternal mRNAs during mouse oogenesis. We report here that by driving the expression of a transgene encoding an Msy2 hairpin dsRNA in growing oocytes using the oocyte-specific Zp3 promoter, the amount of MSY2 protein was reduced by at least 60% in fully grown oocytes. The decrease appeared specific because no decrease was observed in either non-targeted mRNAs or proteins. Fertility of transgenic females was severely reduced. Although transgenic eggs could be inseminated, the eggs did not exhibit the normal series of oscillations in intracellular Ca2+, resume meiosis, undergo cortical granule exocytosis, or ZP2 cleavage to ZP2f. Transgenic oocytes also displayed a higher incidence of both the non-surrounded nucleolus chromatin morphology, and abnormal meiotic spindle formation was observed following oocyte maturation. Transgenic oocytes contained less total mRNA (approximately 75-80% that of non-transgenic oocytes) and displayed a reduced level of protein synthesis. Moreover, several of the maturation-associated changes in protein synthesis failed to occur in the transgenic oocytes. These results support a role for MSY2 in stabilizing maternal mRNAs in growing oocytes, a process essential to generate meiotically and developmentally competent oocytes.

Animals↗
PubMed1996

Editing of the GLuR-B ion channel RNA in vitro by recombinant double-stranded RNA adenosine deaminase.

Double-stranded RNA (dsRNA)-specific adenosine deaminase (DRADA) has been implicated as an enzyme responsible for the editing of RNA transcripts encoding glutamate-gated ion channel subunits (GLuR) in brain. In one case, the editing alters the gene-encoded glutamine (Q) to an arginine (R) located within the channel-forming domain of the alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA) receptor subunit GLuR-B. The result of editing at this site, called the 'Q/R' site, is a profound alteration of the Ca2+ permeability of the GLuR channel. Using recombinantly expressed DRADA proteins, we now demonstrate in vitro that DRADA is indeed involved in editing of the GLuR-B RNA. In addition to the formation of an RNA duplex structure involving exon and intron sequences, Q/R site-selective editing by DRADA also requires a cofactor protein(s) commonly present even in non-neuronal cells. The accuracy and efficiency of this RNA editing system appear to be determined by the quantitative balance between DRADA, cofactor and substrate GLuR-B RNA.

Adenosine Deaminase↗
PubMed2005

Efficient RNA interference in zebrafish embryos using siRNA synthesized with SP6 RNA polymerase.

Double-stranded RNA (dsRNA) has been shown to be a useful tool for silencing genes in zebrafish (Danio rerio), while the blocking specificity of dsRNA is still of major concern for application. It was reported that siRNA (small interfering RNA) prepared by endoribonuclease digestion (esiRNA) could efficiently silence endogenous gene expression in mammalian embryos. To test whether esiRNA could work in zebrafish, we utilized Escherichia coli RNaseIII to digest dsRNA of zebrafish no tail (ntl), a mesoderm determinant in zebrafish and found that esi-ntl could lead to developmental defects, however, the effective dose was so close to the toxic dose that esi-ntl often led to non-specific developmental defects. Consequently, we utilized SP6 RNA polymerase to produce si-ntl, siRNA designed against ntl, by in vitro transcription. By injecting in vitro synthesized si-ntl into zebrafish zygotes, we obtained specific phenocopies of reported mutants of ntl. We achieved up to a 59%no tail phenotype when the injection concentration was as high as 4 microg/microL. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) and whole-mount in situ hybridization analysis showed that si-ntl could largely and specifically reduce mRNA levels of the ntl gene. As a result, our data indicate that esiRNA is unable to cause specific developmental defects in zebrafish, while siRNA should be an alternative for downregulation of specific gene expression in zebrafish in cases where RNAi techniques are applied to zebrafish reverse genetics.

Animals↗
PubMed2005

RDE-2 interacts with MUT-7 to mediate RNA interference in Caenorhabditis elegans.

In Caenorhabditis elegans, the activity of transposable elements is repressed in the germline. One of the mechanisms involved in this repression is RNA interference (RNAi), a process in which dsRNA targets cleavage of mRNAs in a sequence-specific manner. The first gene found to be involved in RNAi and transposon silencing in C.elegans is mut-7, a gene encoding a putative exoribonuclease. Here, we show that the MUT-7 protein resides in complexes of approximately 250 kDa in the nucleus and in the cytosol. In addition, we find that upon triggering of RNAi the cytosolic MUT-7 complex increases in size. This increase is independent of the presence of target RNA, but does depend on the presence of RDE-1 and RDE-4, two proteins involved in small interfering RNA (siRNA) production. Finally, using a yeast two-hybrid screen, we identified RDE-2/MUT-8 as one of the other components of this complex. This protein is encoded by the rde-2/mut-8 locus, previously implicated in RNAi and transposon silencing. Using genetic complementation analysis, we show that the interaction between these two proteins is required for efficient RNAi in vivo. Together these data support a role for the MUT-7/RDE-2 complex downstream of siRNA formation, but upstream of siRNA mediated target RNA recognition, possibly indicating a role in the siRNA amplification step.

ATP-Binding Cassette Transporters↗
PubMed2001

The 90- and 110-kDa human NFAR proteins are translated from two differentially spliced mRNAs encoded on chromosome 19p13.

The NFAR gene (nuclear factor associated with dsRNA) encodes a putative transcription-associated factor that we have shown is a substrate for the interferon-inducible, dsRNA-dependent protein kinase, PKR. However, our protein expression analysis has revealed that NFAR exists as two major protein species of 90 kDa (NFAR-1) and 110 kDa (NFAR-2) in the cell. To resolve the genetic identity of NFAR-1 and -2, we carried out sequence analysis of genomic and cDNA NFAR clones and determined that the coding region of this gene spans 16.2 kb and comprises 21 exons. Our data indicate that NFAR-1 and -2 arise from a single gene on chromosome 19p13 and are generated through alternative splicing events. NFAR-1 (HGMW-approved symbol ILF3) was found to comprise 1 extra exon, 18, that contains several stop codons to ensure termination of the protein at amino acid 702. In contrast, NFAR-2 lacks this exon, though it comprises an additional 3 coding exons (exons 19, 20, and 21) located at the carboxyl region to generate an extended product of 894 amino acids. Our studies, the first to elucidate the gene structure and chromosomal assignment of NFAR, establish the genetic basis for future NFAR research in humans.

Alternative Splicing↗
PubMed1997

Protein P7 of phage phi6 RNA polymerase complex, acquiring of RNA packaging activity by in vitro assembly of the purified protein onto deficient particles.

The RNA polymerase complex of double-stranded RNA bacteriophage phi6 is composed of four proteins, P1, P2, P4 and P7. These four proteins are capable of performing all the functions required for the replication of the double-stranded RNAs of the phi6 genome. The polymerase complex containing the three genomic dsRNA segments is the core particle of the phi6 virion. In this study purified protein P7 was found to form highly asymmetric dimers. Using polyclonal anti-P7 antibody, P7 was shown to be accessible on the surface of the nucleocapsid. Treatment of nucleocapsids with polyclonal anti-P7 antibody released coat protein P8 with ensuing activation of the plus strand RNA synthesis from the resulting core particles. Purified P7 could be assembled onto particles lacking P7 and particles lacking both P2 (RNA polymerase) and P7. In both cases RNA packaging activity was acquired. Assembly of P7 onto deficient particles took place also in the absence of host proteins. Protein P7 is known to be necessary for stable packaging of the three genomic phi6 plus strand RNAs into preformed polymerase complex particles. Additionally, protein P7 seems to be involved in the regulation of plus strand synthesis (i.e. transcription) as a fidelity factor. Particles lacking protein P7 produce anomalous size transcripts. Analysis of the polymerase complex stability revealed that proteins P2, P4 and P7 are independently associated with the major structural protein P1. The number of P7 molecules in one virion was estimated to be 60 and a location at the 5-fold symmetry position is proposed.

Bacteriophage phi 6↗
PubMed1996

Structure and regulation of the Salmonella typhimurium rnc-era-recO operon.

The Escherichia coli rnc-era-recO operon encodes ribonuclease III (RNase III; a dsRNA endonuclease involved in rRNA and mRNA processing and decay), Era (an essential G-protein of unknown functions and RecO (involved in the RecF homologous recombination pathway). Expression of the rnc and era genes is negatively autoregulated: RNase III cleaves the rncO 'operator' in the untranslated leader, destabilizing the operon mRNA. As part of a larger effort to understand RNase III and Era structure and function, we characterized rnc operon structure, function and regulation in the closely related bacterium Salmonella typhimurium. Construction of a S typhimurium strain conditionally defective for RNase III and Era expression showed that Era is essential for cell growth. This mutant strain also enabled selection of recombinant clones containing the intact S typhimurium rnc-era-recO operon, whose nucleotide sequence, predicted protein sequence, and predicted rncO RNA secondary structure were all highly conserved with those of E coli. Furthermore, genetic and biochemical analysis revealed that S typhimurium rnc gene expression is negatively autoregulated by a mechanism very similar or identical to that in E coli, and that the cleavage specificities of RNase IIIs.t. and RNase IIIE.c. are indistinguishable with regard to rncO cleavage and S typhimurium 23S rRNA fragmentation in vivo.

Bacterial Proteins↗
PubMed2006

An unusual Dicer-like1 protein fuels the RNA interference pathway in Trypanosoma brucei.

RNA interference (RNAi) is an evolutionarily conserved gene-silencing pathway that is triggered by double-stranded RNA (dsRNA). Central to this pathway are two ribonucleases: Dicer, a multidomain RNase III family enzyme that initiates RNAi by generating small interfering RNAs (siRNAs), and Argonaute or Slicer, an RNase H signature enzyme that affects cleavage of mRNA. Previous studies in the early diverging protozoan Trypanosoma brucei have established a key role for Argonaute 1 in RNAi. However, the identity of Dicer has not been resolved. Here, we report the identification and functional characterization of a T. brucei Dicer-like enzyme (TbDcl1). Using genetic and biochemical approaches, we provide evidence that TbDcl1 is required for the generation of siRNA-size molecules and for RNAi. Whereas Dicer and Dicer-like proteins are endowed with two adjacent RNase III domains at the carboxyl terminus (RNase IIIa and RNase IIIb), the arrangement of these two domains is unusual in TbDcl1. RNase IIIa is close to the amino terminus, and RNase IIIb is located approximately in the center of the molecule. This domain organization is specific to trypanosomatids and further illustrates the variable structures of protozoan Dicer-like proteins as compared to fungal and metazoan Dicer.

Amino Acid Sequence↗
PubMed2003

[RNA interference in three ES cell lines from different mouse strains].

RNA interference phenomenon in three different murine ES cell lines (MESPU13, B3, and R1) is reported. A vector(pdsGFP) was used that transcribed hairpin double-stranded RNA of GFP gene to transfect ES cells by using lipofectin. The transient transcription of dsRNA induced RNAi (RNA interference) in the ES cells. That is, the double-stranded RNA of GFP gene potently turned down the expression of the GFP gene. On the hand, the linearized plasmid pdsGFP-puro was electroporated into MESPU13 ES cells, and the expression level of GFP after puromycin screening was turned down obviously in about 30% ES cell clones; and in a few clones, the expression level of GFP was not observed under the fluorescence microscope and GFP mRNA was not detectable by RT-PCR. Further more, another vector (pdsOCT4) was constructed that transcribed double-stranded RNA of OCT-4 gene which is specifically expressed in ES cells. ES cell clones that stably integrated the vector were screened after the electrotransfection of the cells with the above construct. 51 random-selected clones were amplified and 48 of them were checked by semi-quantitative RT-PCR. In 11 of them the mRNA of OCT-4 was undetectable by RT-PCR. This means that RNAi can be used to study mammal and human gene's function in ES cell lines from different strain mice.

Animals↗