Mutations in the p53 tumor suppressor are common in the
|
|
- Wilfrid Washington
- 5 years ago
- Views:
Transcription
1 High metastatic potential in mice inheriting a targeted p53 missense mutation Geng Liu*, Timothy J. McDonnell, Roberto Montes de Oca Luna*, Mini Kapoor*, Betsy Mims*, Adel K. El-Naggar, and Guillermina Lozano* Departments of *Molecular Genetics, Molecular Pathology, and Pathology, University of Texas M. D. Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, TX Edited by Peter K. Vogt, The Scripps Research Institute, La Jolla, CA, and approved January 5, 2000 (received for review September 22, 1999) To understand the relevance of p53 missense mutations in vivo,we generated a mouse containing an arg-to-his substitution at p53 amino acid 172, which corresponds to the R175H hot-spot mutation in human tumors by homologous recombination. Inadvertently, this mouse contains the additional deletion of a G nucleotide at a splice junction that attenuates levels of mutant p53 to near wild-type levels. Mice heterozygous for the mutant allele differed from p53 mice in tumor spectrum, with a significant increase in the number of carcinomas and a slight decrease in the number of lymphomas. More importantly, the osteosarcomas and carcinomas that developed in these mutant mice frequently metastasized (69% and 40%, respectively). In contrast, metastasis is rare in osteosarcomas of p53 mice. Loss of heterozygosity studies of tumors indicated loss of heterozygosity in only 1 of 11 tumors. These data indicate clear differences between a p53 missense mutation and a null allele in tumorigenesis in vivo and suggest that the p53r172h g mutant represents a gain-of-function allele. Mutations in the p53 tumor suppressor are common in the etiology of many tumors and disrupt the ability of p53 to induce growth arrest or apoptosis. In particular, the R175H mutation corresponds to one of six hot spot mutations in p53 and accounts for 6% of the missense mutations identified in human cancers (1). Mutant p53 proteins containing missense mutations have been studied in various assays, leading to suggestions that some may yield gain-of-function or dominant negative phenotypes. For example, expression of mutant p53 in p53 null cells yields tumors in syngeneic mice (2) and makes established cells highly tumorigenic (3). When assayed in comparison to other mutants, the p53r175h mutant was always the most detrimental (4 7). The p53r175h mutant cooperates better than any other mutant in ras-mediated transformation (4, 5). Cells expressing the p53r175h mutant are the most tumorigenic, as measured by growth in soft agar (6). In another experiment, genomic instability was measured in fibroblasts derived from Li-Fraumeni syndrome patients heterozygous for the p53r175h mutant (8). These cells exhibited a disrupted spindle checkpoint control and accumulated a polyploid DNA content as compared with cells with other p53 missense mutations, which arrest with a 4n DNA content. These data, coupled with the fact that most p53 mutations ( 80%) occurring in human tumors are missense mutations, suggest that cells with p53 missense mutations have a growth advantage. These in vitro experiments, however, cannot assay the effect of mutant p53 on tumor growth rate, survival, and tumor spectrum in vivo. The generation of mice null for p53 (9, 10) has yielded a plethora of data as to the importance of p53 in vivo. Mice heterozygous or homozygous for a p53 null allele are predisposed to multiple tumor types such as lymphomas, sarcomas, and other tumors resembling to some extent the types of tumors seen in the Li-Fraumeni syndrome. Thymocytes from mice lacking p53 are resistant to cell death by DNA-damaging agents compared with cells from normal mice (11). However, the null mouse model may be inadequate in that the most common alteration in p53 both in somatic cells (1) and in the germline of patients with Li-Fraumeni syndrome is a missense mutation. Thus, it is possible that the existing model has shed light only on the tumorigenic effect of those p53 mutations that are loss of function. A mouse that contains a missense mutation in one of the endogenous p53 genes would be invaluable to understanding p53 missense mutations in vivo. To examine the in vivo significance of p53 missense mutations, we generated a mouse containing an arg-to-his substitution at amino acid 172 (equivalent to the 175 arg-to-his in human) in one of the endogenous p53 alleles by using homologous recombination. Mice with this mutation express low levels of mutant p53. Mice heterozygous for the R172H mutation exhibit differences in tumor spectrum and an increased rate of metastasis as compared with p53 mice. Importantly, most tumors from heterozygous mice do not show loss of heterozygosity (LOH). This allele thus exhibits a gain-of-function phenotype in vivo. Materials and Methods Gene Targeting and Generating the p53r172h g Mice. A 15.3-kb NotI-NotI p53 fragment was isolated from a murine 129 Sv genomic library and was subcloned. The double replacement procedure was used essentially as described (12). The targeting vector for the first targeting step contained the 2.4-kb HindIII- BamHI fragment of p53 intron 1, the PGKneoNTRtkpA cassette (12) (a gift from R. Jaenisch, Massachusetts Institute of Technology), and the 0.9-kb partial PstI-PstI fragment containing p53 intron 6 through intron 9. The resulting plasmid Pgl6 was linearized with SalI and was electroporated into the AB-1 embryonic stem (ES) cells. DNA was harvested, was digested by EcoRI, and was subjected to mini-southern blot analysis (13). One correctly targeted ES cell, 3C12, was expanded and used for the second step of targeting. The targeting plasmid contained the 15.3-kb p53 genomic fragment with the R172H mutation in exon 5. Site-directed PCR mutagenesis protocol (14) was utilized to generate the p53 arg-to-his mutation (CGC to CAC) at codon 172. The PCR product containing the mutation was cloned and sequenced to confirm the desired mutation. The resulting targeting plasmid (36N6) was linearized and electroporated into ES cell clone 3C12. Single strand conformation polymorphism (SSCP) analysis was performed on DNA from ES clones by using a pair of primers flanking the p53 exon 5-intron 5 region: A3 (5 -TAC TCT CCT CCC CTC AAT AAG C-3 ) and A4 (5 -AAT TAC AGA CCT CGG GTG GC-3 ) as described (15). The PCR to detect the TK gene was performed by using the TK This paper was submitted directly (Track II) to the PNAS office. Abbreviations: ES cell, embryonic stem cell; SSCP, single strand conformation polymorphism; LOH, loss of heterozygosity. Present address: Department of Microbiology and Immunology, Universidad Autonoma de Nuevo Leon, Monterrey, NL Mexico. To whom reprint requests should be addressed. gglozano@notes.mdacc.tmc.edu. The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C solely to indicate this fact PNAS April 11, 2000 vol. 97 no. 8
2 GENETICS Fig. 1. The generation of a mouse containing a p53r172h substitution. (a) The genomic organization of the p53 gene and the targeting vectors are shown. The first recombination step results in incorporation of the neo and tk genes (PGKneoNTRtkpA) at the p53 locus. In the second step, the neo and tk genes are replaced with the genomic p53 containing a point mutation (asterisk), resulting in an arg-to-his substitution at amino acid 172. g represents the deletion of a G nucleotide at the intron 2 splice acceptor site. (b) Southern hybridization of EcoRI-digested DNA from several ES cell clones electroporated with the PGKneoNTRtkpA targeting vector. The 17-kb EcoRI fragment represents the wild-type p53 allele whereas the 3.4- and 12-kb EcoRI fragments are the expected sizes for the mutant. The ES cell clone (3C12) in lane 2 is correctly targeted. (c) SSCP analysis using primers spanning the mutation. Lanes: 1, plasmid containing wild-type p53 genomic sequences; 2, plasmid containing mutant p53 genomic sequences; 3, genomic DNA from targeted ES cell clone; 4, genomic DNA from normal ES cells. *, wild-type bands; **, mutant-specific bands. (d) Southern blot hybridization of EcoRI-digested DNA from derivatives of 3C12 cells targeted with a construct containing a missense mutation in p53. Lanes: 1, normal ES cell DNA; 2, 3C12 DNA; 3, DNA from a correctly targeted ES cell clone. primer (5 -ACT GAA GGC TTT ACT ATT GC-3 ) and A6 (5 -CCT CTG TAG CAT GGG CAT CC-3 ). The DNA from targeted ES cell clones were digested with EcoRI and were subjected to the same Southern blot analysis performed in the first step targeting to check the integrity of the locus. Mutant ES cell lines were used for injection into C57 BL6 blastocysts to generate chimeric mice. High percentage chimeras from clone 3C10 were crossed with C57 BL6 female mice and contributed to the germline of mice. The resulting heterozygous mice were either intercrossed to generate heterozygous mice and homozygous mice or were crossed back with C57 BL6 to generate heterozygous mice. Mice in the tumorigenic study came from both crosses. Genotyping of p53r172h g mice was performed by using an allele-specific primer reaction. Primers 172W (5 -CAC ATG ACG GAG GTC GTG ACA CG-3 ) and exon 7 (5 -ATG GTG GTA TAC TCA GAG CC-3 ) amplified the wild-type p53 allele and primers 172M (5 -CAC ATG ACG GAG GTC GTG ACA CA-3 ), and exon 7 amplified the R172H mutant allele in separate reactions. Immunoprecipitation and Reverse Transcription PCR. Mouse embryonic fibroblasts were obtained from dissecting and mincing 12.5-day embryos individually and were maintained in DMEM containing 15% fetal bovine serum. Mouse embryonic fibroblasts were labeled and proteins immunoprecipitated with p53 antibody PAb421 as described (16). Total RNA was isolated from mouse embryonic fibroblast cells homozygous for a p53r172h g allele by using a RNAzol B solution (Tel-Test, Friendswood, TX). Forty micrograms of RNA were used to purify mrna by using Dynabeads (Dynal, Liu et al. PNAS April 11, 2000 vol. 97 no
3 Great Neck, NY) according to the manufacturer s instructions. First-strand cdna was synthesized by using either a polydt primer or p53 primer E11. Using the first-strand cdna as substrate, primers E1a (5 -GAG TTC ATT GGG ACC ATC CTG-3 ) and E11 (5 -AGG ATT GTG TCT CAG CCC TGA-3 ) were used to generate full-length p53 cdna for sequencing analysis. Primers E2 (5 -AGCCAGGAGACATTTTCACC-3 ) and E4 (5 -GCAGGAGCTCCTGACACT CGG-3 ) were also used to clone different splicing variants in the mouse embryonic fibroblasts that were homozygous for the p53r172h g allele. Statistical Analysis. The binomial distribution test for comparing two proportions was used to analyze the statistical significance of the tumor spectrum difference between p53r172h g mice and two groups of p53 mice. LOH Analysis. DNA was isolated from paraffin-embedded tumor samples identified by a pathologist using laser-capture technology. PCR amplification using primers A3 and A4 resulted in a product 384 base pairs long. Although it can be difficult to generate PCR products of this length from paraffin-extracted DNA, we were able to analyze eight of the samples. All samples were also analyzed by using primers A3 and mie5r (5 -CCA TCA CCA TCG GAG CAG CG-3 ), which generated a PCR product of 184 base pairs. Both primer sets gave identical results. PCR products were generated by using touchdown thermocycling conditions: initial denaturation at 95 C for 5 min followed by 10 cycles of denaturation at 94 C for 1 min, annealing at 65 C for 1 min (decreasing 1 C per min until 56 C), and extension at 72 C for 1 min. This was followed by 35 cycles at 94 C for 1 min, 55 C for 1 min, and 72 C for 1 min and final extension at 72 C for 10 min. The PCR reaction was supplemented with [ - 32 P]dCTP, and 5 l of the labeled products were denatured and separated on a 0.5 MDE (FMC) polyacrylamide gel. Electrophoresis was performed for 16 hours at 4.5 W. Gels were dried and bands were visualized by autoradiography. The quantitative in vitro reconstitutional test was performed by SSCP analysis with mutant DNA (representing an LOH tumor sample) mixed with increasing proportions of heterozygous genomic DNA (representing a potential contaminating source) and quantifying the intensities of the mutant and wild-type bands. Results Generation of p53r172h g Mutant Mice. To examine the in vivo significance of p53 missense mutations, we generated a mouse containing an arg-to-his substitution at amino acid 172 (equivalent to the 175 arg-to-his in human) in one of the endogenous p53 alleles by using homologous recombination. The doublereplacement method (12) was used to introduce the mutation into ES cells (Fig. 1a). In brief, the first electroporation with a targeting vector containing the neo and TK genes between p53 homologous sequences marked the locus of interest. The first round of selection occurred in G418, and the G418-resistant ES cell colonies were screened by Southern blotting for the correct targeting event (Fig. 1b). The targeted clone 3C12 was expanded and used for electroporation with a second plasmid containing the p53 gene with the R172H mutation (Fig. 1a). To generate a missense mutation at amino acid 172, a PCR-based site-directed mutagenesis protocol was used. A CGC to CAC substitution at codon 172 was confirmed by sequencing; no other changes were observed. Selection against TK was performed in the presence of 1-(2-deoxy-2-fluoro- -D-arabinofuranosyl)-s-iodouracil (FIAU). In the case of a homologous recombination event, the neo and TK genes are lost and the p53 gene is reconstituted. To identify the targeted missense mutation in the ES cells, we performed SSCP (Fig. 1c) and sequencing analyses to confirm the mutation in the p53 locus. In addition, the restriction digests of the locus confirmed that the p53 gene was reconstituted (Fig. Fig. 2. (a) Analysis of p53 protein levels in fibroblasts from embryos null for p53 (lane 1), wild-type (WT; lanes 2 and 3), or homozygous for the p53r172h g mutation (lane 4). Wild-type fibroblasts were treated with UV to show induction of p53 and confirm position on the gel (lane 3). (b) The structure of the p53 gene around exons 2 4 and the spliced products identified in fibroblasts from homozygous p53r172h g mutant mice. denotes the G nucleotide in intron 2 that is deleted. 1d). Two ES cell clones were obtained, and both were used in blastocyst injections. One gave rise to chimeric mice, which resulted in germline transmission of the allele with the p53r172h mutation. Mice homozygous and heterozygous for the p53 mutant allele were established. Fibroblasts from embryos with these genotypes were also established in culture. Immunoprecipitation analysis with a p53 antibody, PAb421, indicated that homozygous mutant cells expressed levels of the p53r172h mutant protein comparable to wild-type p53 (Fig. 2a). This was not expected because the corresponding mutation in human p53 results in an increased half-life and thus a higher level of protein (5). Detailed analysis by reverse transcription PCR revealed the presence of several p53 mrnas from cells homozygous for the mutant allele, and sequencing indicated three mrna products: full length, deletion of exon 3, and deletion of a part of exon 3 with the aberrant splicing products being the major species (Fig. 2b). Sequencing of genomic DNA in this region revealed deletion of a G nucleotide at the splice acceptor site of intron 2 (Fig. 2b). The incorrectly spliced mrna species result in a frameshift of the p53 coding sequence. This deletion explains both the presence of multiple mrna transcripts and the low levels of mutant protein being produced. Additional experiments performed to determine whether smaller proteins were made by this allele showed the absence of smaller p53 protein products. The deletion occurred in the last cloning step generated by the process of cloning in Escherichia coli because only the targeting construct had this mutation. We named this p53 mutant allele p53r172h g. Analysis of Survival, Tumor Spectrum, and Metastasis in p53r172h g Mice. Because mice were making mutant p53 protein, albeit at a reduced level, we examined a cohort of mice homozygous and heterozygous for the p53r172h g mutation. There were no differences between the survival curves when homozygous and heterozygous p53r172h g mice were compared with p53 and p53 mice of the same mixed C57 BL6 and 129 Sv Liu et al.
4 Table 1. Tumor spectrum of p53r172h g mice Tumor types No. Metastasis Sarcomas Osteosarcomas Fibrosarcomas 10 0 Angiosarcomas 3 0 Lyposarcomas 1 0 Carcinomas Adenocarcinomas 5 2 Hepatocellular carcinomas 4 0 Poorly differentiated carcinomas 1 1 Squamous cell carcinomas 5 2 Islet cell carcinomas 1 1 Lymphoma 9 0 Adenoma 1 0 In a group of 51 p53r172h g/ mice, one or more tumors had developed in 43. A total of 56 tumors were detected and used to analyze the tumor distribution. Fig. 3. Comparisons of survival between p53 null mice and p53r172h g mice. The p53 mice survival data set is from our lab, and the p53 mice survival data set is from Harvey et al. (17). (a) Survival curves of 80 p53r172h g R172H g mice and 40 p53 mice. (b) Survival curves of 92 p53r172h g mice and 150 p53 mice. background (10, 17) (Fig. 3). Like p53 mice, the p53r172h g homozygous mice were highly susceptible to early onset spontaneous tumor formation. The predominant tumor type that developed in these mice (over 75%) was malignant thymic lymphoma. In contrast, the tumor spectrum in the p53r172h g heterozygous mice differed significantly from that of p53 mice (Table 1). Even though the p53r172h g heterozygous mice developed sarcomas at a frequency similar to p53 mice [54 versus 57% (10, 17)], carcinomas were significantly more frequent in the p53r172h g mice than in the p53 mice [29% vs. 10% (17) and 11% (10); P and P 0.025, respectively]. The lymphoma incidence was slightly lower in the p53r172h g mice than in two independent studies of p53 null mice [16% vs. 25% (10) and 32% (17); P 0.25 and P 0.025, respectively]. It is also interesting to note that the latencies of the major tumor types that developed in p53r172h g mice were very similar (Fig. 4). In addition to differences in tumor spectrum, tumors in the p53r172h g heterozygous animals demonstrated a high metastatic potential, especially the osteosarcomas and carcinomas (Fig. 5). In 43 of 51 mice that had tumors, 17 had obvious metastasis nodules in the lung, liver, or both. Eleven of sixteen (69%) osteosarcomas had metastasized. Adenocarcinomas and squamous cell carcinomas also metastasized at high frequency, both at 40%. One unusual tumor, an islet cell carcinoma, also metastasized. Interestingly, none of 10 fibrosarcomas exhibited evidence of metastatic lesions. In contrast, several studies have reported no metastasis (17) or a very low rate of metastasis (10%) (18) from the osteosarcomas in the p53 mice of the same genetic background. There have been no reports of metastasis from carcinomas in p53 mice. Additionally, in our laboratory, we have not seen metastasis in p53 mice nor in p53 null mice with varying copies of the mdm2 null allele (19), suggesting that laboratory conditions per se do not alter this phenotype. Loss of Heterozygosity. Loss of heterozygosity is a hallmark for loss of function of tumor suppressor genes in tumorigenesis. Because the data presented above suggested a gain-of-function activity for the p53r172h g allele, we checked the status of the wild-type p53 allele in different types of tumors from the p53r172h g mice. SSCP analysis can readily distinguish the wild-type and mutant alleles (see Fig. 1c). The in situ lasercapture procedure was used to isolate tumor sample from adjacent normal tissue. In addition, quantitative in vitro reconstitution experiments demonstrated that even up to 15% of Fig. 4. Tumor latency of different tumor types from 56 tumors that developed in 51 p53r172h g mice. Os, osteosarcomas; Fs, fibrosarcomas; As, angiosarcomas; Lp, lyposarcomas; Ad, adenocarcinomas; HC, hepatocellular carcinomas; Pa, poorly differentiated carcinomas; SC, squamous cell carcinomas; IlC, islet cell carcinomas; Ly, lymphomas; Aa, adenoma. GENETICS Liu et al. PNAS April 11, 2000 vol. 97 no
5 a b Fig. 6. A loss of heterozygosity study of tumors from p53r172h g mice. SSCP analysis was performed on tumor samples obtained by using laser capture technology. (a) LOH analysis of DNA from osteosarcomas from mice numbered 395 and 565 using A3 and A4 primers (see Materials and Methods). 565N indicates DNA isolated from normal tissue adjacent to tumor of mouse 565. (b) LOH analysis of DNA from a carcinoma from mouse 3124 using primers A3 and Mie5R (see Materials and Methods). Tail DNA from wild type ( ), homozygous ( g g), and heterozygous ( g ) mice were used as controls to distinguish the two alleles. c normal cell contamination would not obscure the LOH results (data not shown). The LOH results from 11 different tumor samples are shown in Fig. 6 and are summarized in Table 2. Ten of eleven tumors retained the wild-type allele and thus demonstrated no LOH at the p53 locus. Only one adenocarcinoma showed LOH (Fig. 6b). These data suggest that the loss of the wild-type p53 allele is not essential for tumor development and further supports the gain-of-function phenotype of mice inheriting the p53r172h g allele. Discussion p53r172h g mice have a targeted p53 missense mutation, an arg-to-his substitution at amino acid 172, and an additional splicing mutation in intron 2. This allele expressed a small amount of p53 mutant protein. The tumor spectrum of mice heterozygous for this mutation differed from that of p53 Table 2. Summary of LOH results from 11 tumors that developed from p53r172h g/ mice by SSCP analysis Mouse no. Age, weeks Tumor LOH Fig. 5. Frequency of metastasis from tumors of p53r172h g mice. (a) Metastatic lesions from an osteosarcoma that developed in a p53r172h g mouse. (A) Liver. (B) Lung. (b) Representative histologies of tumors and their metastatic spread. A, C, and E represent primary tumors; B, D, F, and G represent their corresponding metastases. (A and B) Hepatocelluar carcinomas with metastasis in lung. (C and D) Lung adenocarcinomas with metastasis in myocardium (E, F, and G) Osteosarcoma with metastasis in liver (F) and lung (G). (c) Frequency of metastasis from p53r172h g mice vs. p53 mice. p53 metastasis data are from Harvey et al. (17) and Tervana et al. (18), respectively Osteosarcoma Adjacent normal tissue Osteosarcoma Fibrosarcoma Fibrosarcoma Adenocarcinoma Adenocarcinoma Hepatocellular carcinoma Adjacent normal tissue Squamous cell carcinoma Squamous cell carcinoma Lymphoma Lymphoma, LOH;, no LOH Liu et al.
6 mice, as did the metastatic potential. Thus, we have established mice with a p53 mutant allele by homologous recombination. The differences in tumor spectrum and the increased frequency of metastasis in the p53r172h g mice suggest that this allele represents a gain-of-function. Unlike transgenic models used to study the functional significance of overexpression of p53 missense mutations in vivo (20 22), this mouse generated by homologous recombination expressed only a small amount of protein. Therefore, the observed phenotypes most likely represent the intrinsic properties of the mutant protein. The increased incidence of metastasis in p53r172h g mice does not appear to be caused by differences in tumor spectrum. Increased metastasis is observed in osteosarcomas of p53r172h g mice as compared with p53 mice, yet the numbers of sarcomas arising in both genotypes are identical (10, 17). Previous studies using cell lines lacking p53 or expressing a mutant p53 showed that those cells expressing mutant p53 readily metastasized to the lung of syngeneic mice (23), further supporting the gain-of-function phenotype of p53 missense mutations. Another important observation is that the fibrosarcomas as opposed to osteosarcomas and carcinomas that develop in p53r172h g mice did not metastasize. Of 10 fibrosarcomas that were identified, none metastasized, whereas of 16 osteosarcomas, 11 did. In contrast, Nf-2 heterozygous mice showed a high frequency of metastasis from both osteosarcomas and fibrosarcomas (24). This finding suggests that the determinants of metastasis are different for different tissue types. LOH studies were also performed on 11 different tumors from p53r172h g mice to determine whether the wild-type p53 allele had been lost. Only 1 of 11 tumors showed loss of heterozygosity, suggesting that loss of wild-type p53 was not important to tumor development. This is in contrast to LOH studies in p53 mice, in which 50% of the mice under 18 months of age were shown to have LOH at the wild-type p53 allele whereas 85% of mice over 18 months of age showed no LOH (25). The age of the mice when tumors were diagnosed varied in our experiments and did not seem to have an impact on loss of the wild-type allele. These data indicate a gain-of-function phenotype for the p53r172h g allele. The mutant protein may disrupt other cellular processes in the cell that cannot be compensated for by the presence of wild-type p53. Alternatively, the data may also suggest that this allele functions as a dominant negative. The mutant protein has the ability to form tetramers with the wild-type p53, effectively removing it from the cell. Surprisingly, even though differences in tumor spectrum and metastasis are observed, there was no significant difference in the survival of these mice. It is possible that other alterations were rate limiting in mice or that, for osteosarcomas and carcinomas, metastasis is not a contributing factor to survival in mice. The mating of p53r172h g mice to other tumor models will help address some of these issues. Our study clearly establishes that the p53r172h mutant represents a gain-of-function allele. These data support clinical studies showing a high correlation of this specific p53 mutation, an arg-to-his substitution at amino acid 175, and extremely poor prognosis in humans (26). We thank Larry Donehower for the p53 data set, Howard Thames for help with statistical analysis, and Lisa Amelse for help with the mice. This work was supported by grants from the National Institutes of Health. 1. Greenblatt, M. S., Bennett, W. P., Hollstein, M. & Harris, C. C. (1994) Cancer Res. 54, Wolf, D., Harris, N. & Rotter, V. (1984) Cell 38, Eliyahu, D., Michalovitz, D. & Oren, M. (1985) Nature (London) 316, Finlay, C. A., Hinds, P. W. & Levine, A. J. (1989) Cell 57, Hinds, P. W., Finlay, C. A., Quartin, R. S., Baker, S. J., Fearon, E. R., Vogelstein, B. & Levine, A. J. (1990) Cell Growth Differ. 1, Dittmer, D., Pati, S., Zambetti, G., Chu, S., Teresky, A. K., Moore, M., Finlay, C. & Levine, A. J. (1993) Nat. Genet. 4, Slingerland, J. M., Jenkins, J. R. & Benchimol, S. (1993) EMBO J. 12, Gualberto, A., Aldape, K., Kozakiewicz, K. & Tlsty, T. D. (1998) Proc. Natl. Acad. Sci. USA 95, Donehower, L. A., Harvey, M., Slagle, B. L., McArthur, M. J., Montgomery, C. A., Jr., Butel, J. S. & Bradley, A. (1992) Nature (London) 356, Jacks, T., Remington, L., Williams, B. O., Schmitt, E. M., Halachmi, S., Bronson, R. T. & Weinberg, R. A. (1994) Curr. Biol. 4, Lowe, S. W., Schmitt, E. M., Smith, S. W., Osborne, B. A. & Jacks, T. (1993) Nature (London) 362, Wu, H., Liu, X. & Jaenisch, R. (1994) Proc. Natl. Acad. Sci. USA 91, McMahon, A. P. & Bradley, A. (1990) Cell 62, Haiguchi, R. (1990) Recombinant PCR Protocols: 1990 (Academic, San Diego). 15. Evans, S. C., Mims, B., McMasters, K. M., Foster, C. J., deandrade, M., Amos, C. I., Strong, L. C. & Lozano, G. (1998) Hum. Genet. 102, Kapoor, M. & Lozano, G. (1998) Proc. Natl. Acad. Sci. USA 95, Harvey, M., McArthur, M. J., Montgomery, C. A., Jr., Butel, J. S., Bradley, A. & Donehower, L. A. (1993) Nat. Genet. 5, Taverna, D., Ullman-Cullere, M., Rayburn, H., Bronson, R. T. & Hynes, R. O. (1998) Cancer Res. 58, McDonnell, T., Montes De Oca Luna, R., Cho, S., Amelse, L. L., Chavez- Reyes, A. & Lozano, G. (1999) J. Pathol. 188, Lavigueur, A., Maltby, V., Mock, D., Rossant, J., Pawson, T. & Bernstein, A. (1989) Mol. Cell. Biol. 9, Wang, X. J., Greenhalgh, D. A., Jiang, A., He, D., Zhong, L., Medina, D., Brinkley, B. R. & Roop, D. R. (1998) Oncogene 17, Li, B., Murphy, K. L., Laucirica, R., Kittrell, F., Medina, D. & Rosen, J. M. (1998) Oncogene 16, Pohl, J., Goldfinger, N., Radler-Pohl, A., Rotter, V. & Schirrmacher, V. (1988) Mol. Cell. Biol. 8, McClatchey, A. I., Saotome, I., Mercer, K., Crowley, D., Gusella, J. F., Bronson, R. T. & Jacks, T. (1998) Genes Dev. 12, Venkatachalam, S., Shi, Y. P., Jones, S. N., Vogel, H., Bradley, A., Pinkel, D. & Donehower, L. A. (1998) EMBO J. 17, Goh, H. S., Yao, J. & Smith, D. R. (1995) Cancer Res. 55, GENETICS Liu et al. PNAS April 11, 2000 vol. 97 no
Supplementary Document
Supplementary Document 1. Supplementary Table legends 2. Supplementary Figure legends 3. Supplementary Tables 4. Supplementary Figures 5. Supplementary References 1. Supplementary Table legends Suppl.
More informationc Tuj1(-) apoptotic live 1 DIV 2 DIV 1 DIV 2 DIV Tuj1(+) Tuj1/GFP/DAPI Tuj1 DAPI GFP
Supplementary Figure 1 Establishment of the gain- and loss-of-function experiments and cell survival assays. a Relative expression of mature mir-484 30 20 10 0 **** **** NCP mir- 484P NCP mir- 484P b Relative
More informationSupplementary Appendix
Supplementary Appendix This appendix has been provided by the authors to give readers additional information about their work. Supplement to: Sherman SI, Wirth LJ, Droz J-P, et al. Motesanib diphosphate
More informationa) Primary cultures derived from the pancreas of an 11-week-old Pdx1-Cre; K-MADM-p53
1 2 3 4 5 6 7 8 9 10 Supplementary Figure 1. Induction of p53 LOH by MADM. a) Primary cultures derived from the pancreas of an 11-week-old Pdx1-Cre; K-MADM-p53 mouse revealed increased p53 KO/KO (green,
More informationBeta Thalassemia Case Study Introduction to Bioinformatics
Beta Thalassemia Case Study Sami Khuri Department of Computer Science San José State University San José, California, USA sami.khuri@sjsu.edu www.cs.sjsu.edu/faculty/khuri Outline v Hemoglobin v Alpha
More informationNature Structural & Molecular Biology: doi: /nsmb Supplementary Figure 1
Supplementary Figure 1 U1 inhibition causes a shift of RNA-seq reads from exons to introns. (a) Evidence for the high purity of 4-shU-labeled RNAs used for RNA-seq. HeLa cells transfected with control
More informationSupplementary Table 3. 3 UTR primer sequences. Primer sequences used to amplify and clone the 3 UTR of each indicated gene are listed.
Supplemental Figure 1. DLKI-DIO3 mirna/mrna complementarity. Complementarity between the indicated DLK1-DIO3 cluster mirnas and the UTR of SOX2, SOX9, HIF1A, ZEB1, ZEB2, STAT3 and CDH1with mirsvr and PhastCons
More informationSupplemental Data. Shin et al. Plant Cell. (2012) /tpc YFP N
MYC YFP N PIF5 YFP C N-TIC TIC Supplemental Data. Shin et al. Plant Cell. ()..5/tpc..95 Supplemental Figure. TIC interacts with MYC in the nucleus. Bimolecular fluorescence complementation assay using
More informationFigure S1. Analysis of genomic and cdna sequences of the targeted regions in WT-KI and
Figure S1. Analysis of genomic and sequences of the targeted regions in and indicated mutant KI cells, with WT and corresponding mutant sequences underlined. (A) cells; (B) K21E-KI cells; (C) D33A-KI cells;
More informationBIOLOGY 621 Identification of the Snorks
Name: Date: Block: BIOLOGY 621 Identification of the Snorks INTRODUCTION: In this simulation activity, you will examine the DNA sequence of a fictitious organism - the Snork. Snorks were discovered on
More informationAbbreviations: P- paraffin-embedded section; C, cryosection; Bio-SA, biotin-streptavidin-conjugated fluorescein amplification.
Supplementary Table 1. Sequence of primers for real time PCR. Gene Forward primer Reverse primer S25 5 -GTG GTC CAC ACT ACT CTC TGA GTT TC-3 5 - GAC TTT CCG GCA TCC TTC TTC-3 Mafa cds 5 -CTT CAG CAA GGA
More informationSupplementary Figure 1 MicroRNA expression in human synovial fibroblasts from different locations. MicroRNA, which were identified by RNAseq as most
Supplementary Figure 1 MicroRNA expression in human synovial fibroblasts from different locations. MicroRNA, which were identified by RNAseq as most differentially expressed between human synovial fibroblasts
More informationSupplementary Materials
Supplementary Materials 1 Supplementary Table 1. List of primers used for quantitative PCR analysis. Gene name Gene symbol Accession IDs Sequence range Product Primer sequences size (bp) β-actin Actb gi
More informationSupplementary Figure 1 a
Supplementary Figure a Normalized expression/tbp (A.U.).6... Trip-br transcripts Trans Trans Trans b..5. Trip-br Ctrl LPS Normalized expression/tbp (A.U.) c Trip-br transcripts. adipocytes.... Trans Trans
More informationCANCER. Inherited Cancer Syndromes. Affects 25% of US population. Kills 19% of US population (2nd largest killer after heart disease)
CANCER Affects 25% of US population Kills 19% of US population (2nd largest killer after heart disease) NOT one disease but 200-300 different defects Etiologic Factors In Cancer: Relative contributions
More informationSupplementary Figure 1. ROS induces rapid Sod1 nuclear localization in a dosagedependent manner. WT yeast cells (SZy1051) were treated with 4NQO at
Supplementary Figure 1. ROS induces rapid Sod1 nuclear localization in a dosagedependent manner. WT yeast cells (SZy1051) were treated with 4NQO at different concentrations for 30 min and analyzed for
More informationBeta Thalassemia Sami Khuri Department of Computer Science San José State University Spring 2015
Bioinformatics in Medical Product Development SMPD 287 Three Beta Thalassemia Sami Khuri Department of Computer Science San José State University Hemoglobin Outline Anatomy of a gene Hemoglobinopathies
More informationTITLE: A Mouse Model to Investigate the Role of DBC2 in Breast Cancer
AD Award Number: W81XWH-04-1-0325 TITLE: A Mouse Model to Investigate the Role of DBC2 in Breast Cancer PRINCIPAL INVESTIGATOR: Valerie Boka CONTRACTING ORGANIZATION: University of Texas Health Science
More information*To whom correspondence should be addressed. This PDF file includes:
www.sciencemag.org/cgi/content/full/science.1212182/dc1 Supporting Online Material for Partial Retraction to Detection of an Infectious Retrovirus, XMRV, in Blood Cells of Patients with Chronic Fatigue
More informationCulture Density (OD600) 0.1. Culture Density (OD600) Culture Density (OD600) Culture Density (OD600) Culture Density (OD600)
A. B. C. D. E. PA JSRI JSRI 2 PA DSAM DSAM 2 DSAM 3 PA LNAP LNAP 2 LNAP 3 PAO Fcor Fcor 2 Fcor 3 PAO Wtho Wtho 2 Wtho 3 Wtho 4 DTSB Low Iron 2 4 6 8 2 4 6 8 2 22 DTSB Low Iron 2 4 6 8 2 4 6 8 2 22 DTSB
More informationMultistep nature of cancer development. Cancer genes
Multistep nature of cancer development Phenotypic progression loss of control over cell growth/death (neoplasm) invasiveness (carcinoma) distal spread (metastatic tumor) Genetic progression multiple genetic
More informationTumor suppressor genes D R. S H O S S E I N I - A S L
Tumor suppressor genes 1 D R. S H O S S E I N I - A S L What is a Tumor Suppressor Gene? 2 A tumor suppressor gene is a type of cancer gene that is created by loss-of function mutations. In contrast to
More informationSUPPLEMENTARY INFORMATION
doi: 10.1038/nature05883 SUPPLEMENTARY INFORMATION Supplemental Figure 1 Prostaglandin agonists and antagonists alter runx1/cmyb expression. a-e, Embryos were exposed to (b) PGE2 and (c) PGI2 (20μM) and
More information(Stratagene, La Jolla, CA) (Supplemental Fig. 1A). A 5.4-kb EcoRI fragment
SUPPLEMENTAL INFORMATION Supplemental Methods Generation of RyR2-S2808D Mice Murine genomic RyR2 clones were isolated from a 129/SvEvTacfBR λ-phage library (Stratagene, La Jolla, CA) (Supplemental Fig.
More informationTable S1. Oligonucleotides used for the in-house RT-PCR assays targeting the M, H7 or N9. Assay (s) Target Name Sequence (5 3 ) Comments
SUPPLEMENTAL INFORMATION 2 3 Table S. Oligonucleotides used for the in-house RT-PCR assays targeting the M, H7 or N9 genes. Assay (s) Target Name Sequence (5 3 ) Comments CDC M InfA Forward (NS), CDC M
More informationWhat we know about Li-Fraumeni syndrome
What we know about Li-Fraumeni syndrome Dr Helen Hanson Consultant in Cancer Genetics St Georges Hospital, South-West Thames Regional Genetics Service History of LFS 1969 Li and Fraumeni describe four
More informationMutation Screening and Association Studies of the Human UCP 3 Gene in Normoglycemic and NIDDM Morbidly Obese Patients
Mutation Screening and Association Studies of the Human UCP 3 Gene in Normoglycemic and NIDDM Morbidly Obese Patients Shuichi OTABE, Karine CLEMENT, Séverine DUBOIS, Frederic LEPRETRE, Veronique PELLOUX,
More informationThe autoimmune disease-associated PTPN22 variant promotes calpain-mediated Lyp/Pep
SUPPLEMENTARY INFORMATION The autoimmune disease-associated PTPN22 variant promotes calpain-mediated Lyp/Pep degradation associated with lymphocyte and dendritic cell hyperresponsiveness Jinyi Zhang, Naima
More informationToluidin-Staining of mast cells Ear tissue was fixed with Carnoy (60% ethanol, 30% chloroform, 10% acetic acid) overnight at 4 C, afterwards
Toluidin-Staining of mast cells Ear tissue was fixed with Carnoy (60% ethanol, 30% chloroform, 10% acetic acid) overnight at 4 C, afterwards incubated in 100 % ethanol overnight at 4 C and embedded in
More informationwithout LOI phenotype by breeding female wild-type C57BL/6J and male H19 +/.
Sakatani et al. 1 Supporting Online Material Materials and methods Mice and genotyping: H19 mutant mice with C57BL/6J background carrying a deletion in the structural H19 gene (3 kb) and 10 kb of 5 flanking
More informationIntegration Solutions
Integration Solutions (1) a) With no active glycosyltransferase of either type, an ii individual would not be able to add any sugars to the O form of the lipopolysaccharide. Thus, the only lipopolysaccharide
More informationA smart acid nanosystem for ultrasensitive. live cell mrna imaging by the target-triggered intracellular self-assembly
Electronic Supplementary Material (ESI) for Chemical Science. This journal is The Royal Society of Chemistry 2017 A smart ZnO@polydopamine-nucleic acid nanosystem for ultrasensitive live cell mrna imaging
More informationSupplementary Figures
Supplementary Figures Supplementary Figure 1. H3F3B expression in lung cancer. a. Comparison of H3F3B expression in relapsed and non-relapsed lung cancer patients. b. Prognosis of two groups of lung cancer
More informationSupplementary Figure 1
Supplementary Figure 1 3 3 3 1 1 Bregma -1.6mm 3 : Bregma Ref) Http://www.mbl.org/atlas165/atlas165_start.html Bregma -.18mm Supplementary Figure 1 Schematic representation of the utilized brain slice
More informationCitation for published version (APA): Oosterveer, M. H. (2009). Control of metabolic flux by nutrient sensors Groningen: s.n.
University of Groningen Control of metabolic flux by nutrient sensors Oosterveer, Maaike IMPORTANT NOTE: You are advised to consult the publisher's version (publisher's PDF) if you wish to cite from it.
More informationPhylogenetic analysis of human and chicken importins. Only five of six importins were studied because
Supplementary Figure S1 Phylogenetic analysis of human and chicken importins. Only five of six importins were studied because importin-α6 was shown to be testis-specific. Human and chicken importin protein
More informationCD31 5'-AGA GAC GGT CTT GTC GCA GT-3' 5 ' -TAC TGG GCT TCG AGA GCA GT-3'
Table S1. The primer sets used for real-time RT-PCR analysis. Gene Forward Reverse VEGF PDGFB TGF-β MCP-1 5'-GTT GCA GCA TGA ATC TGA GG-3' 5'-GGA GAC TCT TCG AGG AGC ACT T-3' 5'-GAA TCA GGC ATC GAG AGA
More informationWhat do you think of when you here the word genome?
What do you think of when you here the word genome? What do you think of when you here the word genome? Personal Genomics Outline Review of pre-lab work Genomics and Medicine Case Overview & Assignment
More informationMUTATIONS, MUTAGENESIS, AND CARCINOGENESIS. (Start your clickers)
MUTATIONS, MUTAGENESIS, AND CARCINOGENESIS (Start your clickers) How do mutations arise? And how do they affect a cell and its organism? Mutations: heritable changes in genes Mutations occur in DNA But
More informationNucleotide Sequence of the Australian Bluetongue Virus Serotype 1 RNA Segment 10
J. gen. Virol. (1988), 69, 945-949. Printed in Great Britain 945 Key words: BTV/genome segment lo/nucleotide sequence Nucleotide Sequence of the Australian Bluetongue Virus Serotype 1 RNA Segment 10 By
More informationSupplementary Materials and Methods
DD2 suppresses tumorigenicity of ovarian cancer cells by limiting cancer stem cell population Chunhua Han et al. Supplementary Materials and Methods Analysis of publicly available datasets: To analyze
More informationSupplementary Table 2. Conserved regulatory elements in the promoters of CD36.
Supplementary Table 1. RT-qPCR primers for CD3, PPARg and CEBP. Assay Forward Primer Reverse Primer 1A CAT TTG TGG CCT TGT GCT CTT TGA TGA GTC ACA GAA AGA ATC AAT TC 1B AGG AAA TGA ACT GAT GAG TCA CAG
More informationComputational Systems Biology: Biology X
Bud Mishra Room 1002, 715 Broadway, Courant Institute, NYU, New York, USA L#4:(October-0-4-2010) Cancer and Signals 1 2 1 2 Evidence in Favor Somatic mutations, Aneuploidy, Copy-number changes and LOH
More informationProblem Set 8 Key 1 of 8
7.06 2003 Problem Set 8 Key 1 of 8 7.06 2003 Problem Set 8 Key 1. As a bright MD/PhD, you are interested in questions about the control of cell number in the body. Recently, you've seen three patients
More informationCh 7 Mutation. A heritable change in DNA Random Source of genetic variation in a species may be advantageous, deleterious, neutral
Ch 7 Mutation A heritable change in DNA Random Source of genetic variation in a species may be advantageous, deleterious, neutral Mutation (+ sexual reproduction) + natural selection = evolution Types
More informationCharacterizing intra-host influenza virus populations to predict emergence
Characterizing intra-host influenza virus populations to predict emergence June 12, 2012 Forum on Microbial Threats Washington, DC Elodie Ghedin Center for Vaccine Research Dept. Computational & Systems
More informationSingle-Molecule Analysis of Gene Expression Using Two-Color RNA- Labeling in Live Yeast
Supplemental Figures, Tables and Results Single-Molecule Analysis of Gene Expression Using Two-Color RNA- Labeling in Live Yeast Sami Hocine 1, Pascal Raymond 2, Daniel Zenklusen 2, Jeffrey A. Chao 1 &
More informationNucleic Acid Testing - Oncology. Molecular Diagnosis. Gain/Loss of Nucleic Acid. Objectives. MYCN and Neuroblastoma. Molecular Diagnosis
Nucleic Acid Testing - Oncology Molecular Diagnosis Nucleic acid based testing in Oncology Gross alterations in DNA content of tumors (ploidy) Gain/Loss of nucleic acids Markers of Clonality Oncogene/Tumor
More informationMolecular Diagnosis. Nucleic acid based testing in Oncology
Molecular Diagnosis Nucleic acid based testing in Oncology Objectives Describe uses of NAT in Oncology Diagnosis, Prediction, monitoring. Genetics Screening, presymptomatic testing, diagnostic testing,
More informationSupplemental Information. Th17 Lymphocytes Induce Neuronal. Cell Death in a Human ipsc-based. Model of Parkinson's Disease
Cell Stem Cell, Volume 23 Supplemental Information Th17 Lymphocytes Induce Neuronal Cell Death in a Human ipsc-based Model of Parkinson's Disease Annika Sommer, Franz Maxreiter, Florian Krach, Tanja Fadler,
More informationEarly Embryonic Development
Early Embryonic Development Maternal effect gene products set the stage by controlling the expression of the first embryonic genes. 1. Transcription factors 2. Receptors 3. Regulatory proteins Maternal
More informationDetection of 549 new HLA alleles in potential stem cell donors from the United States, Poland and Germany
HLA ISSN 2059-2302 BRIEF COMMUNICATION Detection of 549 new HLA alleles in potential stem cell donors from the United States, Poland and Germany C. J. Hernández-Frederick 1, N. Cereb 2,A.S.Giani 1, J.
More informationSupplementary Figure 1. AdipoR1 silencing and overexpression controls. (a) Representative blots (upper and lower panels) showing the AdipoR1 protein
Supplementary Figure 1. AdipoR1 silencing and overexpression controls. (a) Representative blots (upper and lower panels) showing the AdipoR1 protein content relative to GAPDH in two independent experiments.
More informationSUPPLEMENTARY DATA. Supplementary Table 1. Primer sequences for qrt-pcr
Supplementary Table 1. Primer sequences for qrt-pcr Gene PRDM16 UCP1 PGC1α Dio2 Elovl3 Cidea Cox8b PPARγ AP2 mttfam CyCs Nampt NRF1 16s-rRNA Hexokinase 2, intron 9 β-actin Primer Sequences 5'-CCA CCA GCG
More informationSALSA MLPA KIT P060-B2 SMA
SALSA MLPA KIT P6-B2 SMA Lot 111, 511: As compared to the previous version B1 (lot 11), the 88 and 96 nt DNA Denaturation control fragments have been replaced (QDX2). Please note that, in contrast to the
More informationSupplemental Information. Cancer-Associated Fibroblasts Neutralize. the Anti-tumor Effect of CSF1 Receptor Blockade
Cancer Cell, Volume 32 Supplemental Information Cancer-Associated Fibroblasts Neutralize the Anti-tumor Effect of CSF1 Receptor Blockade by Inducing PMN-MDSC Infiltration of Tumors Vinit Kumar, Laxminarasimha
More informationStudying The Role Of DNA Mismatch Repair In Brain Cancer Malignancy
Kavya Puchhalapalli CALS Honors Project Report Spring 2017 Studying The Role Of DNA Mismatch Repair In Brain Cancer Malignancy Abstract Malignant brain tumors including medulloblastomas and primitive neuroectodermal
More informationAllelic and Haplotype Frequencies of the p53 Polymorphisms in Brain Tumor Patients
Physiol. Res. 51: 59-64, 2002 Allelic and Haplotype Frequencies of the p53 Polymorphisms in Brain Tumor Patients E. BIROŠ, I. KALINA, A. KOHÚT 1, E. BOGYIOVÁ 2, J. ŠALAGOVIČ, I. ŠULLA 3 Department of Medical
More informationSupplemental Data Mutant p53 Gain of Function in Two Mouse Models of Li-Fraumeni Syndrome
Supplemental Data Mutant p53 Gain of Function in Two Mouse Models of Li-Fraumeni Syndrome S1 Kenneth P. Olive, David A. Tuveson, Zachary C. Ruhe, Bob Yin, Nicholas A. Willis, Roderick T. Bronson, Denise
More informationDescription of Supplementary Files. File Name: Supplementary Information Description: Supplementary Figures and Supplementary Tables
Description of Supplementary Files File Name: Supplementary Information Description: Supplementary Figures and Supplementary Tables Supplementary Figure 1: (A), HCT116 IDH1-WT and IDH1-R132H cells were
More informationCancer Genetics 204 (2011) 45e52
Cancer Genetics 204 (2011) 45e52 Exon scanning by reverse transcriptaseepolymerase chain reaction for detection of known and novel EML4eALK fusion variants in nonesmall cell lung cancer Heather R. Sanders
More informationMRC-Holland MLPA. Description version 19;
SALSA MLPA probemix P6-B2 SMA Lot B2-712, B2-312, B2-111, B2-511: As compared to the previous version B1 (lot B1-11), the 88 and 96 nt DNA Denaturation control fragments have been replaced (QDX2). SPINAL
More informationChapter 12-4 DNA Mutations Notes
Chapter 12-4 DNA Mutations Notes I. Mutations Introduction A. Definition: Changes in the DNA sequence that affect genetic information B. Mutagen= physical or chemical agent that interacts with DNA to cause
More informationSupplementary Figure 1
Supplementary Figure 1 Supplementary Figure 1. Lats1/2 deleted ihbs and ihps showed decreased transcripts of hepatocyte related genes (a and b) Western blots (a) and recombination PCR (b) of control and
More informationTHE UMD TP53 MUTATION DATABASE UPDATES AND BENEFITS. Pr. Thierry Soussi
THE UMD TP53 MUTATION DATABASE UPDATES AND BENEFITS Pr. Thierry Soussi thierry.soussi@ki.se thierry.soussi@upmc.fr TP53: 33 YEARS AND COUNTING STRUCTURE FUNCTION RELATIONSHIP OF WILD AND MUTANT TP53 1984
More informationwww.lessonplansinc.com Topic: Protein Synthesis - Sentence Activity Summary: Students will simulate transcription and translation by building a sentence/polypeptide from words/amino acids. Goals & Objectives:
More informationPlasmids Western blot analysis and immunostaining Flow Cytometry Cell surface biotinylation RNA isolation and cdna synthesis
Plasmids psuper-retro-s100a10 shrna1 was constructed by cloning the dsdna oligo 5 -GAT CCC CGT GGG CTT CCA GAG CTT CTT TCA AGA GAA GAA GCT CTG GAA GCC CAC TTT TTA-3 and 5 -AGC TTA AAA AGT GGG CTT CCA GAG
More informationMutational analysis of p53 gene in sporadic breast carcinoma
Pak. J. Biochem. Mol. Biol. 2015; 48(3): 79-83 Mutational analysis of p53 gene in sporadic breast carcinoma Irsa Mateen and Saba Irshad* Institute of Biochemistry and Biotechnology, University of the Punjab,
More informationA Rapid and Sensitive Chip-based Assay for Detection of rpob Gene Mutations Conferring Rifampicin Resistance in Mycobacterium tuberculosis (TB).
A Rapid and Sensitive Chip-based Assay for Detection of rpob Gene Mutations Conferring Rifampicin Resistance in Mycobacterium tuberculosis (TB). Wanyuan Ao, Steve Aldous, Evelyn Woodruff, Brian Hicke,
More informationSUPPLEMENTARY INFORMATION Glucosylceramide synthase (GlcT-1) in the fat body controls energy metabolism in Drosophila
SUPPLEMENTARY INFORMATION Glucosylceramide synthase (GlcT-1) in the fat body controls energy metabolism in Drosophila Ayako Kohyama-Koganeya, 1,2 Takuji Nabetani, 1 Masayuki Miura, 2,3 Yoshio Hirabayashi
More informationNature Immunology: doi: /ni.3836
Supplementary Figure 1 Recombinant LIGHT-VTP induces pericyte contractility and endothelial cell activation. (a) Western blot showing purification steps for full length murine LIGHT-VTP (CGKRK) protein:
More informationInsulin Resistance. Biol 405 Molecular Medicine
Insulin Resistance Biol 405 Molecular Medicine Insulin resistance: a subnormal biological response to insulin. Defects of either insulin secretion or insulin action can cause diabetes mellitus. Insulin-dependent
More informationAn oncogenic form of p53 confers a dominant, gain-of-function phenotype that disrupts spindle checkpoint control
Proc. Natl. Acad. Sci. USA Vol. 95, pp. 5166 5171, April 1998 Genetics An oncogenic form of p53 confers a dominant, gain-of-function phenotype that disrupts spindle checkpoint control ANTONIO GUALBERTO*,
More informationMRC-Holland MLPA. Description version 06; 23 December 2016
SALSA MLPA probemix P417-B2 BAP1 Lot B2-1216. As compared to version B1 (lot B1-0215), two reference probes have been added and two target probes have a minor change in length. The BAP1 (BRCA1 associated
More informationMedium-Chain Acyl-CoA Dehydrogenase (MCAD) splicing mutations identified in newborns with an abnormal MS/MS profile
EURASNET Workshop on RNA splicing and genetic diagnosis, London, UK Medium-Chain Acyl-CoA Dehydrogenase (MCAD) splicing mutations identified in newborns with an abnormal MS/MS profile Brage Storstein Andresen
More informationMEDICAL GENOMICS LABORATORY. Next-Gen Sequencing and Deletion/Duplication Analysis of NF1 Only (NF1-NG)
Next-Gen Sequencing and Deletion/Duplication Analysis of NF1 Only (NF1-NG) Ordering Information Acceptable specimen types: Fresh blood sample (3-6 ml EDTA; no time limitations associated with receipt)
More informationSupplementary Figure 1a
Supplementary Figure 1a Hours: E-cadherin TGF-β On TGF-β Off 0 12 24 36 48 24 48 72 Vimentin βactin Fig. S1a. Treatment of AML12 cells with TGF-β induces EMT. Treatment of AML12 cells with TGF-β results
More informationConstruction of a hepatocellular carcinoma cell line that stably expresses stathmin with a Ser25 phosphorylation site mutation
Construction of a hepatocellular carcinoma cell line that stably expresses stathmin with a Ser25 phosphorylation site mutation J. Du 1, Z.H. Tao 2, J. Li 2, Y.K. Liu 3 and L. Gan 2 1 Department of Chemistry,
More informationIdentification of a Mouse Cytomegalovirus Gene Selectively Targeting CD86 Expression on Antigen-Presenting Cells
JOURNAL OF VIROLOGY, Dec. 2004, p. 13062 13071 Vol. 78, No. 23 0022-538X/04/$08.00 0 DOI: 10.1128/JVI.78.23.13062 13071.2004 Copyright 2004, American Society for Microbiology. All Rights Reserved. Identification
More information(a) Schematic diagram of the FS mutation of UVRAG in exon 8 containing the highly instable
Supplementary Figure 1. Frameshift (FS) mutation in UVRAG. (a) Schematic diagram of the FS mutation of UVRAG in exon 8 containing the highly instable A 10 DNA repeat, generating a premature stop codon
More informationRetention of wild-type p53 in tumors from p53 heterozygous mice: reduction of p53 dosage can promote cancer formation
The EMBO Journal Vol.17 No.16 pp.4657 4667, 1998 Retention of wild-type p53 in tumors from p53 heterozygous mice: reduction of p53 dosage can promote cancer formation Sundaresan Venkatachalam 1, Yu-Ping
More informationCell Biology and Cancer
Name: Cell Biology and Cancer Date: Questions 1. BRCA1 and BRCA2 are what types of genes? 2. List two ways that cancerous and healthy cells differ. 3. Which organelle makes proteins? 4. At what phase of
More informationTITLE: The Role of hcdc4 as a Tumor Suppressor Gene in Genomic Instability Underlying Prostate Cancer
AD Award Number: TITLE: The Role of hcdc4 as a Tumor Suppressor Gene in Genomic Instability Underlying Prostate Cancer PRINCIPAL INVESTIGATOR: Audrey van Drogen, Ph.D. CONTRACTING ORGANIZATION: Sidney
More informationBacterial Gene Finding CMSC 423
Bacterial Gene Finding CMSC 423 Finding Signals in DNA We just have a long string of A, C, G, Ts. How can we find the signals encoded in it? Suppose you encountered a language you didn t know. How would
More informationMRC-Holland MLPA. Description version 30; 06 June 2017
SALSA MLPA probemix P081-C1/P082-C1 NF1 P081 Lot C1-0517, C1-0114. As compared to the previous B2 version (lot B2-0813, B2-0912), 11 target probes are replaced or added, and 10 new reference probes are
More informationFigure S1. Generation of inducible PTEN deficient mice and the BMMCs (A) B6.129 Pten loxp/loxp mice were mated with B6.
Figure S1. Generation of inducible PTEN deficient mice and the BMMCs (A) B6.129 Pten loxp/loxp mice were mated with B6.129-Gt(ROSA)26Sor tm1(cre/ert2)tyj /J mice. To induce deletion of the Pten locus,
More informationHST.161 Molecular Biology and Genetics in Modern Medicine Fall 2007
MIT OpenCourseWare http://ocw.mit.edu HST.161 Molecular Biology and Genetics in Modern Medicine Fall 2007 For information about citing these materials or our Terms of Use, visit: http://ocw.mit.edu/terms.
More informationMolecular Pathology of Ovarian Carcinoma with Morphological Correlation
Molecular athology of Ovarian Carcinoma with Morphological Correlation Kathleen R. Cho, M.D. Comprehensive Cancer Center and Departments of athology and Internal Medicine University of Michigan Medical
More informationMRC-Holland MLPA. Description version 29; 31 July 2015
SALSA MLPA probemix P081-C1/P082-C1 NF1 P081 Lot C1-0114. As compared to the previous B2 version (lot 0813 and 0912), 11 target probes are replaced or added, and 10 new reference probes are included. P082
More informationoncogenes-and- tumour-suppressor-genes)
Special topics in tumor biochemistry oncogenes-and- tumour-suppressor-genes) Speaker: Prof. Jiunn-Jye Chuu E-Mail: jjchuu@mail.stust.edu.tw Genetic Basis of Cancer Cancer-causing mutations Disease of aging
More informationMalignant Amelanotic Melanoma of the Pleura without Primary Skin Lesion: An Autopsy Case Report. a a*
2009 63 6 379 384 Malignant Amelanotic Melanoma of the Pleura without Primary Skin Lesion: An Autopsy Case Report a b a a a* a b 380 63 6 Chest x ray and computed tomography (CT). A, Chest x ray on admission
More informationBIOL2005 WORKSHEET 2008
BIOL2005 WORKSHEET 2008 Answer all 6 questions in the space provided using additional sheets where necessary. Hand your completed answers in to the Biology office by 3 p.m. Friday 8th February. 1. Your
More informationof TERT, MLL4, CCNE1, SENP5, and ROCK1 on tumor development were discussed.
Supplementary Note The potential association and implications of HBV integration at known and putative cancer genes of TERT, MLL4, CCNE1, SENP5, and ROCK1 on tumor development were discussed. Human telomerase
More informationLezione 10. Sommario. Bioinformatica. Lezione 10: Sintesi proteica Synthesis of proteins Central dogma: DNA makes RNA makes proteins Genetic code
Lezione 10 Bioinformatica Mauro Ceccanti e Alberto Paoluzzi Lezione 10: Sintesi proteica Synthesis of proteins Dip. Informatica e Automazione Università Roma Tre Dip. Medicina Clinica Università La Sapienza
More informationFrequency of mosaicism points towards mutation prone early cleavage cell divisions.
Frequency of mosaicism points towards mutation prone early cleavage cell divisions. Chad Harland, Wouter Coppieters, Latifa Karim, Carole Charlier, Michel Georges Germ-line de novo mutations Definition:
More informationBHP 2-7 and Nthy-ori 3-1 cells were grown in RPMI1640 medium (Hyclone) supplemented with 10% fetal bovine serum (Gibco), 2mM L-glutamine, and 100 U/mL
1 2 3 4 Materials and Methods Cell culture BHP 2-7 and Nthy-ori 3-1 cells were grown in RPMI1640 medium (Hyclone) 5 supplemented with 10% fetal bovine serum (Gibco), 2mM L-glutamine, and 100 U/mL 6 penicillin-streptomycin.
More informationReceived 29 December 1998/Accepted 9 March 1999
JOURNAL OF VIROLOGY, June 1999, p. 4794 4805 Vol. 73, No. 6 0022-538X/99/$04.00 0 Copyright 1999, American Society for Microbiology. All Rights Reserved. Molecular Requirements for Human Immunodeficiency
More informationMolecular Characterization of the NF2 Gene in Korean Patients with Neurofibromatosis Type 2: A Report of Four Novel Mutations
Korean J Lab Med 2010;30:190-4 DOI 10.3343/kjlm.2010.30.2.190 Original Article Diagnostic Genetics Molecular Characterization of the NF2 Gene in Korean Patients with Neurofibromatosis Type 2: A Report
More informationChromothripsis: A New Mechanism For Tumorigenesis? i Fellow s Conference Cheryl Carlson 6/10/2011
Chromothripsis: A New Mechanism For Tumorigenesis? i Fellow s Conference Cheryl Carlson 6/10/2011 Massive Genomic Rearrangement Acquired in a Single Catastrophic Event during Cancer Development Cell 144,
More informationSupplementary Figure 1
Metastatic melanoma Primary melanoma Healthy human skin Supplementary Figure 1 CD22 IgG4 Supplementary Figure 1: Immunohisochemical analysis of CD22+ (left) and IgG4 (right), cells (shown in red and indicated
More information