[目的]本研究将FASTmrEMMA、最小角回归(least angle regression,LARS)和随机森林(random forest,RF)方法应用于全基因组选择,以提高植物数量性状预测的准确性和效率,为植物遗传和育种提供有益信息。[方法]对拟南芥自然群体的模拟数据...[目的]本研究将FASTmrEMMA、最小角回归(least angle regression,LARS)和随机森林(random forest,RF)方法应用于全基因组选择,以提高植物数量性状预测的准确性和效率,为植物遗传和育种提供有益信息。[方法]对拟南芥自然群体的模拟数据和真实数据进行全基因组预测。在模拟数据分析中,设置不同的表型缺失率,以平均绝对误差(mean absolute error,MAE)、均方误差(mean squared error,MSE)、预测模型拟合度和计算时间为指标,比较基于最小角回归和随机森林的两阶段算法(two-stage algorithm based on least angle regression and random forest,TSLRF)、基于随机森林的两阶段变量选择(two-stage stepwise variable selection based on random forest,TSRF)、随机森林和全基因组最佳线性无偏预测(genomic best linear unbiased prediction,GBLUP)4种方法的优劣。在拟南芥真实数据研究中,针对长日照花期(days to flowering under long day,LD)、春化长日照花期(days to flowering under long day with vernalization,LDV)和短日照花期(days to flowering under short day,SD)实施全基因组预测,并利用这些表型预测值与观测值进行全基因组关联分析,以比较上述4种全基因组选择方法的性能。[结果]模拟研究表明:在不同表型缺失率下,TSLRF的全基因组预测准确度和预测模型拟合度均较高;真实数据的TSLRF分析也获得相似的结论,且检测到40个已报道与目标性状显著关联的基因。[结论]TSLRF方法的全基因组预测准确度和模型拟合度较高,计算速度快,为分子育种和优异亲本组合的预测提供理论依据。展开更多
The development and progression of gastric cancer involves a number of genetic and epigenetic alterations of tumor suppressor and tumor-related genes. The majority of differentiated carcinomas arise from intestinal me...The development and progression of gastric cancer involves a number of genetic and epigenetic alterations of tumor suppressor and tumor-related genes. The majority of differentiated carcinomas arise from intestinal metaplastic mucosa and exhibit structurally altered tumor suppressor genes, typified by p53, which is inactivated via the classic two-hit mechanism, i.e. loss of heterozygosity （LOH） and mutation of the remaining allele. LOH at certain chromosomal loci accumulates during tumor progression. Approximately 20% of differentiated carcinomas show evidence of mutator pathway tumorigenesis due to hMLH1 inactivation via hypermethylation of promoter CpG islands, and exhibit high-frequency microsatellite instability. In contrast, undifferentiated carcinomas rarely exhibit structurally altered tumor suppressor genes. For instance, while methylation of E-cadherin is often observed in undifferentiated carcinomas, mutation of this gene is generally associated with the progression from differentiated to undifferentiated carcinomas. Hypermethylation of tumor suppressor and tumor-related genes, including APC, CHFR, DAP- kinase, DCC, E-cadherin, GSTP1, hMLH1, p16, PTEN, RASSF1A, RUNX3, and TSLC1, can be detected in both differentiated and undifferentiated carcinomas at varying frequencies. However, the significance of the hypermethylation varies according to the analyzed genomic region, and hypermethylation of these genes can also be present in non-neoplastic gastric epithelia. Promoter demethylation of specific genes, such as MAGE and synudein y, can occur during the progressive stages of both histological types, and is associated with patient prognosis. Thus, while the molecular pathways of gastric carcinogenesis are dependent on histological background, specific genetic alterations can still be used for risk assessment, diagnosis, and prognosis.展开更多
Objective To genetically correct a disease-causing point mutation in human induced pluripotent stem cells （iPSCs） derived from a hemophilia B patient. Methods First, the disease-causing mutation was detected by ...Objective To genetically correct a disease-causing point mutation in human induced pluripotent stem cells （iPSCs） derived from a hemophilia B patient. Methods First, the disease-causing mutation was detected by sequencing the encoding area of human coagulation factor IX （F IX） gene. Genomic DNA was extracted from the iPSCs, and the primers were designed to amplify the eight exons of F IX. Next, the point mutation in those iPSCs was genetically corrected using CRISPR/Cas9 technology in the presence of a 129-nucleotide homologous repair template that contained two synonymous mutations. Then, top 8 potential off-target sites were subsequently analyzed using Sanger sequencing. Finally, the corrected clones were differentiated into hepatocyte-like cells, and the secretion of F IX was validated by immunocytochemistry and ELISA assay.Results The cell line bore a missense mutation in the 6th coding exon （c.676 C〉T） of F IX gene. Correction of the point mutation was achieved via CRISPR/Cas9 technology in situ with a high efficacy at about 22% （10/45） and no off-target effects detected in the corrected iPSC clones. F IX secretion, which was further visualized by immunocytochemistry and quantified by ELISA in vitro, reached about 6 ng/ml on day 21 of differentiation procedure. Conclusions Mutations in human disease-specific iPSCs could be precisely corrected by CRISPR/Cas9 technology, and corrected cells still maintained hepatic differentiation capability. Our findings might throw a light on iPSC-based personalized therapies in the clinical application, especially for hemophilia B.展开更多
文摘[目的]本研究将FASTmrEMMA、最小角回归(least angle regression,LARS)和随机森林(random forest,RF)方法应用于全基因组选择,以提高植物数量性状预测的准确性和效率,为植物遗传和育种提供有益信息。[方法]对拟南芥自然群体的模拟数据和真实数据进行全基因组预测。在模拟数据分析中,设置不同的表型缺失率,以平均绝对误差(mean absolute error,MAE)、均方误差(mean squared error,MSE)、预测模型拟合度和计算时间为指标,比较基于最小角回归和随机森林的两阶段算法(two-stage algorithm based on least angle regression and random forest,TSLRF)、基于随机森林的两阶段变量选择(two-stage stepwise variable selection based on random forest,TSRF)、随机森林和全基因组最佳线性无偏预测(genomic best linear unbiased prediction,GBLUP)4种方法的优劣。在拟南芥真实数据研究中,针对长日照花期(days to flowering under long day,LD)、春化长日照花期(days to flowering under long day with vernalization,LDV)和短日照花期(days to flowering under short day,SD)实施全基因组预测,并利用这些表型预测值与观测值进行全基因组关联分析,以比较上述4种全基因组选择方法的性能。[结果]模拟研究表明:在不同表型缺失率下,TSLRF的全基因组预测准确度和预测模型拟合度均较高;真实数据的TSLRF分析也获得相似的结论,且检测到40个已报道与目标性状显著关联的基因。[结论]TSLRF方法的全基因组预测准确度和模型拟合度较高,计算速度快,为分子育种和优异亲本组合的预测提供理论依据。
文摘The development and progression of gastric cancer involves a number of genetic and epigenetic alterations of tumor suppressor and tumor-related genes. The majority of differentiated carcinomas arise from intestinal metaplastic mucosa and exhibit structurally altered tumor suppressor genes, typified by p53, which is inactivated via the classic two-hit mechanism, i.e. loss of heterozygosity （LOH） and mutation of the remaining allele. LOH at certain chromosomal loci accumulates during tumor progression. Approximately 20% of differentiated carcinomas show evidence of mutator pathway tumorigenesis due to hMLH1 inactivation via hypermethylation of promoter CpG islands, and exhibit high-frequency microsatellite instability. In contrast, undifferentiated carcinomas rarely exhibit structurally altered tumor suppressor genes. For instance, while methylation of E-cadherin is often observed in undifferentiated carcinomas, mutation of this gene is generally associated with the progression from differentiated to undifferentiated carcinomas. Hypermethylation of tumor suppressor and tumor-related genes, including APC, CHFR, DAP- kinase, DCC, E-cadherin, GSTP1, hMLH1, p16, PTEN, RASSF1A, RUNX3, and TSLC1, can be detected in both differentiated and undifferentiated carcinomas at varying frequencies. However, the significance of the hypermethylation varies according to the analyzed genomic region, and hypermethylation of these genes can also be present in non-neoplastic gastric epithelia. Promoter demethylation of specific genes, such as MAGE and synudein y, can occur during the progressive stages of both histological types, and is associated with patient prognosis. Thus, while the molecular pathways of gastric carcinogenesis are dependent on histological background, specific genetic alterations can still be used for risk assessment, diagnosis, and prognosis.
基金Supported by the National Science and Technology Major Project(2011ZX09102-010-04)
文摘Objective To genetically correct a disease-causing point mutation in human induced pluripotent stem cells （iPSCs） derived from a hemophilia B patient. Methods First, the disease-causing mutation was detected by sequencing the encoding area of human coagulation factor IX （F IX） gene. Genomic DNA was extracted from the iPSCs, and the primers were designed to amplify the eight exons of F IX. Next, the point mutation in those iPSCs was genetically corrected using CRISPR/Cas9 technology in the presence of a 129-nucleotide homologous repair template that contained two synonymous mutations. Then, top 8 potential off-target sites were subsequently analyzed using Sanger sequencing. Finally, the corrected clones were differentiated into hepatocyte-like cells, and the secretion of F IX was validated by immunocytochemistry and ELISA assay.Results The cell line bore a missense mutation in the 6th coding exon （c.676 C〉T） of F IX gene. Correction of the point mutation was achieved via CRISPR/Cas9 technology in situ with a high efficacy at about 22% （10/45） and no off-target effects detected in the corrected iPSC clones. F IX secretion, which was further visualized by immunocytochemistry and quantified by ELISA in vitro, reached about 6 ng/ml on day 21 of differentiation procedure. Conclusions Mutations in human disease-specific iPSCs could be precisely corrected by CRISPR/Cas9 technology, and corrected cells still maintained hepatic differentiation capability. Our findings might throw a light on iPSC-based personalized therapies in the clinical application, especially for hemophilia B.
