原发性肝癌P53基因第249密码子突变的研究

用聚合酶链反应－限制性片段长度多态性（ＰＣＲ－ＲＦＬＰ）技术对７例原发性肝细胞癌（ＰＨＣ）进行Ｐ＾５３基因第２４９密码子突变的检测。结果：７例肝细胞癌中３例有Ｐ＾５３基因第２４９密码子突变，突变率达４２．９％。     

Mutational hotspot in the p53 gene in human hepatocellular carcinomas.

Human hepatocellular carcinomas (HCC) from patients in Qidong, an area of high incidence in China, in which both hepatitis B virus and aflatoxin B1 are risk factors, were analysed for mutations in p53, a putative tumour-suppressor gene. Eight of the 16 HCC had a point mutation at the third base position of codon 249. The G----T transversion in seven HCC DNA samples and the G----C transversion in the other HCC are consistent with mutations caused by aflatoxin B1 in mutagenesis experiments. No mutations were found in exons 5,6,8 or the remainder of exon 7. These results contrast with p53 mutations previously reported in carcinomas and sarcomas of human lung, colon, oesophagus and breast; these are primarily scattered over four of the five evolutionarily conserved domains, which include codon 249 (refs 4-9). We suggest that the mutant p53 protein may be responsible for a selective clonal expansion of hepatocytes during carcinogenesis.     

广西肝癌中HBV感染与N-ras基因突变的研究

应用ＰＣＲ－ＳＳＣＰ和免疫组化法检测２９例广西南部肝癌组织中的Ｎ－ｒａｓ基因突变和ＨＢＶ感染状况，结果，肝癌中Ｎ－ｒａｓ基因在第２￣３７密码子之间的突变率为７９．３％，其中２２例有２￣５个突变位点，该基因突变也见于癌旁组织，肝组织中ＨＢｓＡｇ和ＨＢｓＡｇ和ＨＢｘＡｇ检出率分别为８６．２％和７９．３％，两者具有相关性，并与Ｎ－ｒａｓ基因突变率呈相平行的趋势。因广西南部的肝癌与ＡＦＢ１污染有关，本研究     

用计算机对人类TSPYl基因P53结合位点的鉴定

根据p53下游基因在其调节区域（启动子或内含子）含有与P53蛋白特异性结合的一致性序列5’-RRRCWWGYYYN（0-13）RRRCWWGYYY-3’，R—G或A，W—T或A，Y—C或T，N—A，C，T，G。用计算机对人类基因组中P53结合位点进行了研究，发现Y染色体上的TSPY1基因内含子中含有这样的一致性序列5’-GGGCTAGTTTtgGAGCTAGCCT-3’，意味着TSPY1基因有可能是一个p53下游基因。     

Selective G to T mutations of p53 gene in hepatocellular carcinoma from southern Africa.

Hepatocellular carcinoma (HCC) is a prevalent cancer in sub-Saharan Africa and eastern Asia. Hepatitis B virus and aflatoxins are risk factors for HCC, but the molecular mechanism of human hepatocellular carcinogenesis is largely unknown. Abnormalities in the structure and expression of the tumour-suppressor gene p53 are frequent in HCC cell lines, and allelic losses from chromosome 17p have been found in HCCs from China and Japan. Here we report on allelic deletions from chromosome 17p and mutations of the p53 gene found in 50% of primary HCCs from southern Africa. Four of five mutations detected were G----T substitutions, with clustering at codon 249. This mutation specificity could reflect exposure to a specific carcinogen, one candidate being aflatoxin B1 (ref. 7), a food contaminant in Africa, which is both a mutagen that induces G to T substitution and a liver-specific carcinogen.     

黄曲霉毒素高污染区肝细胞癌P53基因高频率定点突变

用聚合酶链反应及限制性片段长度多态性（ＰＣＲ＿ＲＦＬＰ）分析技术，对广西黄曲霉毒素高污染２例肝细胞癌（ＨＣＣ）进行分析，检查Ｐ５３基因第７外显子的２４９密码子的突变频率。结果发现３６／５２例ＨＣＣ中２４８密码子有集中的点突变，频率为６９．２％。Ｐ５３基因突变热点与乙型肝炎病毒感染无关。提示黄曲霉毒素Ｂ１是突变热点的主要原因。     

Mutations analysis of STK_(11) gene in Chinese families with Peutz-Jeghers syndrome

Peutz-Jeghers syndrome (PJS) is an autosomal dominant disease that is characterized by multiple gastrointestinal hamartomatous polyps and melanin spots on lips and buccal mucosa at young age[1,2]. Previous studies have demonstrated that PJS predisposes carriers to cancers of gastrointestinal tract, uterus, ovary, testis, breast and other extragastrointestinal organs[3—5]. The STK11 gene, encoding a serine/threonine kinase at chro-mosome 19p13.3, was identified in 1998 as the main causativ…     

Mutations analysis of STK11 ene in Chinese families with Peutz-Jeghers syndrome

Peutz-Jeghers syndrome (PJS) is an autosomal dominantly inherited disease characterized by multiple gastrointestinal hamartomatous polyps and melanin spots on lips and buccal mucosa, with an increased risk for various cancers. ThePJS gene, a potential tumour suppressor gene, encoding a serine/ threonie kinase (STK11), was mapped to chromosome 19p13.3. To investigate the mutations of STK11 gene in Chinese with PJS, we analyzed its coding sequence in fifteen patientsand twenty unaffected members of six families, including three multigenerational families with PJS and three sporadic families with PJS, by PCR, PCR-DHPLC and DNA sequencing techniques. Ten point mutations were found in the six families, including five missense mutations, one acceptor-splice site mutation, a nonsense mutation and three silent mutations. Our data showed that five missense mutations occurrd at codon 123 (CAG to CAT) in exon 2, codon 161 (ATT to AGT) in exon 4,codon 194 (GAC to GAG) in exon 4, codon 245 (CTC to TTC) in exon 5 and codon 354 (TTC to TTG) in exon 8. One kind of nonsense mutation was detected at codon 37(CAG to TAG) in exon 1. Furthermore, we found an intronic mutation at a splice-acceptor site: a one base substitution from AG to AA in intron 4. These mutations were not detected in 20 normal DNA samples. In three sporadic families, only in one patient, we detected a missense mutation in exon 5. In addition, we found three silent mutations, which may cause polymorphisms of STK11 gene in introns 1(+36), 3(-51) and 5(+27). These results indicated that the point mutation in STK11 might be involved in PJS pathogenesis. Mutation frequency is higher in the families suffering PJS in three or more generations than that of the sporadic cases.     

南宁地区肝细胞性肝癌中突变型p53蛋白表达

用ＡＢＣ免疫组织化学方法，对南宁地区居民中１３例肝细胞性肝癌（ＨＣＣ）的活检组织进行研究。发现其中８例有突变型ｐ＾５３蛋白呈强阳性表达，占ＨＣＣ病例数６４．３％。南宁地区某些县，如扶绥县是我国著名的ＨＣＣ高发区，不但ＨＢＶ感染率高，而且是全国少有的ＡＦＢ１高污染区，后者的作用不能忽视。与江苏启东和南部非洲ＨＣＣ高发区一样，估计也会出现ｐ＾５３基因突变热点。     

人血管生长素基因的化学合成与克隆

采用固相亚磷酸胺法化学合成并克隆人血管生长素基因，该基因全长４２３ｂｐ，其中３６９ｂｐ的结构基因全部选用大肠杆菌偏爱的密码子，在结构基因上游设置了适合原核非融合蛋白表达的核糖体结合位点的Ｓ－Ｄ序列。采用两级退火一步拼接法成功克隆全长基因，该基因适合在多种融合或非融合原核表达载体中表达。     

High frequency of p53 intronic point mutations in laryngeal squamous cell carcinoma

Intronic point mutations are rare and totally unknown for human laryngeal squamous cell carcinoma (LSCC). To explore the relationship of p53 gene intronic mutation to the development of human LSCC, DNA was extracted from both tumor tissues and matched normal tissues of 55 patients with LSCC in northeast of China. Polymerase chain reaction amplification-single strand conformational polymorphism (PCR-SSCP) combined with silver staining and DNA direct sequencing were used to detect mutations in exons 7～8 (p53E7 and p53E8) and introns 7～8 (p53I7 and p53I8) of p53 gene. The p53E7 mutation was detected in 17 out of 55 patients, and the p53I7 mutation in 21 patients. No mutation was found at p53E8 or p53I8 site. The difference between tumor group and paired normal group on the rates of both p53E7 and p53I7 mutations was statistically significant. The rate of p53I7 mutations in tumor tissue was higher than that of normal tissue, and so was that of p53E7. Sequence analysis revealed that most p53I7 mutations were at the nucleotides in the branch point sequence or the polypyrimidine tract in the 3′-splice acceptor site of the intron 7. The high incidence of p53 gene intronic mutation in LSCC indicates that genetic changes within the noncoding region of the p53 gene may serve as an alternative mechanism of activating the pathogenesis of human laryngeal squamous cell carcinoma. Mutations in the noncoding region of this gene should be further studied.     

Germ-line transmission of a mutated p53 gene in a cancer-prone family with Li-Fraumeni syndrome

Tumour suppressor genes, whose usual function seems to be controlling normal cell proliferation, have been implicated in many inherited and sporadic forms of malignancies Much evidence supports the concept of tumour formation by loss-of-function mutations in suppressor genes, as predicted by the two-hit model of Knudson and DeMars. The suppressor gene, p53, is affected in such a manner by numerous mutations, which occur in a variety of human tumours. These mutations usually represent the loss of one allele and the substitution of a single base in the other. We have now analysed the p53 gene in a family affected by Li-Fraumeni syndrome, a rare autosomal dominant syndrome characterized by the occurrence of diverse mesenchymal and epithelial neoplasms at multiple sites. In some instances the neoplasms seem to be related to exposure to carcinogens, including ionizing radiation. The Li-Fraumeni family that we studied had noncancerous skin fibroblasts (NSF) with an unusual radiation-resistant phenotype. DNA derived from the NSF cells of four family members, spanning two generations, had the same point mutation in codon 245 (GGC----GAC) of the p53 gene. This mutation leads to substitution of aspartic acid for glycine in one of the regions identified as a frequent target of point mutations in p53. The NSF cell lines with the mutation also retained the normal p53 allele. This inherited p53 mutation may predispose the members of this family to increased susceptibility to cancer.     

Complete nucleotide sequence of bacteriophage MS2 RNA: primary and secondary structure of the replicase gene.

Bacteriophage MS2 RNA is 3,569 nucleotides long. The nucleotide sequence has been established for the third and last gene, which codes for the replicase protein. A secondary structure model has also been proposed. Biological properties, such as ribosome binding and codon interactions can now be discussed on a molecular basis. As the sequences for the other regions of this RNA have been published already, the complete, primary chemical structure of a viral genome has now been established.     

胃癌中p53基因失活及其蛋白产物的免疫组化分析

应用ＰＣＲ－ＳＳＣＰ银染，Ｓｏｕｔｈｅｒｎ杂我及免疫组化等方法同步研究２５例胃癌标本的ｐ５３基因突变，杂合性丢失及蛋白持表达，在２２例胃癌中同时获得有关ｐ５３基因突变和杂合性丢失的检测结果，被检出ｐ５３基因突变的５例胃癌中，２例伴有杂合性丢失，３例仅有ｐ５３基因突变。     

Evolutionary genomics: codon volatility does not detect selection

Plotkin et al. introduce a method to detect selection that is based on an index called codon volatility and that uses only the sequence of a single genome, claiming that this method is applicable to a large range of sequenced organisms. Volatility for a given codon is the ratio of non-synonymous codons to all sense codons accessible by one point mutation. The significance of each gene's volatility is assessed by comparison with a simulated distribution of 10(6) synonymous versions of each gene, with synonymous codons drawn randomly from average genome frequencies. Here we re-examine their method and data and find that codon volatility does not detect selection, and that, even if it did, the genomes of Mycobacterium tuberculosis and Plasmodium falciparum, as well as those of most sequenced organisms, do not meet the assumptions necessary for application of their method.     

p53抗体与恶性肿瘤

p53基因是人类肿瘤中突变频率最高的抑癌基因,几乎发生于所有的恶性肿瘤.突变基因编码的p53蛋白释放入血,可诱发机体自身免疫应答,产生p53自身抗体.在肿瘤病人和高危人群中检测血清p53抗体可以反映早期p53基因突变,作为一种新的肿瘤生物学指标,p53抗体有望在恶性肿瘤的早期诊断、治疗、预后、监测、复发等方面发挥重要作用.     

淫羊藿苷与过氧化氢酶相互作用的荧光光谱法和分子对接研究

利用过氧化氢酶的内源荧光,通过荧光光谱法对淫羊藿苷与过氧化氢酶的相互作用进行了研究.结果表明,淫羊藿苷主要以静电作用力与过氧化氢酶发生相互作用,16℃时结合常数和结合位点数分别为4.59×104 L/mol和1.00.根据F（o）rster非辐射能量转移理论,计算二者相互作用的结合距离为4.05 nm.对二者之间的相互作用进行了分子模拟,表明淫羊藿苷在过氧化氢酶4个亚基的中心区域通过静电作用力、疏水作用力和氢键作用力与过氧化氢酶发生相互作用,与荧光光谱法的研究结果相一致.     

Detection of a new mutation (1467-A) for the pedigree with mucopolysaccharidosis type Ⅱ from a Chinese family

Mucopolysaccharidosis type Ⅱ is of high genetic heterogeneity. PCR-DNA sequencing was used to study the mutation hot spots in the IDS gene of a Chinese MPS Ⅱ pedigree. A new mutation （1467-A） not yet reported worldwide was detected. This mutation located at 448th codon in the coding region of exon 9 deletes one “A” at the end of 1467 bp （cDNA）. The frame-shift mutation makes the peptide chain shorten from amino acids 550 to 459, probably altering the configuration of IDS enzyme protein remarkably and lowering the activation of IDS greatly. Therefore it is supposed to be the direct cause of the patient with MPS Ⅱ and to be a necessary premise for prenatal gene diagnosis.     

头颈部鳞癌中p53-mdm2基因异常的研究

目的:检测头颈部鳞癌患者p53基因突变、mdm2基因扩增的状况,了解其与头颈部鳞癌患者的性别、鳞癌分级、淋巴结转移等的相关性及两异常基因的相关性。方法:收集50例行手术切除的头颈部鳞癌患者的新鲜肿瘤组织及其相应的癌旁正常组织,提取标本DNA;用PCR-SS-CP-银染法检测p53基因第5~8外显子的突变状况;用dPCR法检测mdm2基因扩增情况;采用SPSS 10.0统计软件包行2检验分析实验结果。结果:在50例头颈部鳞癌患者标本中,检测出17例存在p53基因突变,突变率为34%,所有的癌旁正常组织未发现p53基因突变;在50例鳞癌标本中检测出6例标本存在mdm2基因扩增,扩增率为12%,其中有一例同时存在p53基因突变;p53基因突变、mdm2基因扩增与患者的鳞癌分级、淋巴结转移、性别等相关性分析结果均无统计学意义(P>0.05)。结论:p53基因突变与mdm2基因扩增在头颈部鳞癌中较常见,可能是头颈部鳞癌发生发展的主要分子机制。