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Volume 45, Issue 4
Displaying 1-8 of 8 articles from this issue
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Takao Sekiya2001 Volume 45 Issue 4 Pages 205-214
Published: December 15, 2001
Released on J-STAGE: March 31, 2009
JOURNAL FREE ACCESSWe developed mobility shift analysis of single-stranded DNAs on neutral polyacrylamide gel electrophoresis to detect base changes in DNA. Originally this method follows digestion of genomic DNA with restriction endonucleases, denaturation in alkaline solution, and electrophoresis in a neutral polyacrylamide gel. After transfer to a nylon membrane, the mobility shift due to a nucleotide substitution of a single-stranded DNA fragment could be detected by hybridization with a nick-translated DNA. As the mobility shift caused by nucleotide substitutions might be due to a conformational change of single-stranded DNAs, we designate the features of single-stranded DNAs as single-strand conformation polymorphisms (SSCPs). Invention of the polymerase chain reaction (PCR) made it possible to amplify specific regions of genomic sequences efficiently. The method can simultaneously label amplified DNA fragments by using labeled substrates. After denaturation of PCR products, electrophoresis of obtained single-stranded DNA fragments in nondenaturing polyacrylamide gels and detection of mobility shifts by radioautography can reveal the presence of base substitutions. Most single base changes in up to 300-base fragments could be detected as mobility shifts. Thus the combined use of SSCP and PCR could provide a rapid and sensitive method for the detection of base changes in given sequences of genomic DNA. Many mutations resulted in activation of oncogenes and inactivation of tumor suppressor genes were detected by this PCR-SSCP technique.View full abstractDownload PDF (12472K) -
Masami Muramatsu2001 Volume 45 Issue 4 Pages 215-218
Published: December 15, 2001
Released on J-STAGE: March 31, 2009
JOURNAL FREE ACCESSThe development of modern molecular biology strongly depends on the progress of “electrophoresis” techniques. Since Tiselius, the resolution of electrophoresis separation of proteins had been elevated dramatically, especially after the advent of SDS-polyacrylamide gel by Laemmli, until it became two-dimensional by O'Farrell. On the other hand, nucleic acids became also the target of this technique ended up with the establishment of DNA-sequencing gel by Maxam-Gilbert and Sanger. This technique aided by gene cloning technology has determined a great number of genes with respect to the nucleotide sequence. This also led to the elucidation of the relationship between the structure and the function of genes in eukaryotes as well as prokaryotes. The emerging issue was then to determine the whole genomic as well as mRNA (cDNA) sequences of a species including human in order to understand the riddle of life on the molecular level. The Human Genome Project began in 1989/1990 by the leadership of NIH, USA as a consortium of several countries including U. K., Japan, France and Germany. A venture business Celeara Genomics came in the competition using another procedure named“whole genome shot-gun”. They reconciled in June 2000, by shaking hands at both sides of the President Clinton, announcing that draft sequence of the human genome had been completed. The results were published in February issues of Nature and Science this year (2001). The results were mostly similar confirming their efforts were correct. But, some unexpected results were also obtained; e. g. the unexpectedly low number of human genes, very high rate of appearance of trasposon-type repetitive sequences etc. The author will interpret the results carefully and discuss further on interesting points leading to proteomes.View full abstractDownload PDF (680K) -
Hideki Kambara, Masao Kamahori, Kazunori Okano, Guohua Zou2001 Volume 45 Issue 4 Pages 219-225
Published: December 15, 2001
Released on J-STAGE: March 31, 2009
JOURNAL FREE ACCESSThe human genome project has been successfully carried out and the draft sequence of the human genome has been clarified. The next step is to understand gene functions and to use the genome information in various fields including the biomedical field. As the genetic characteristics are dependent on SNPs, the development of a rapid, inexpensive and high throughput SNP analysis method is required. As the one base difference has to be analyzed in the SNP analysis, the simple hybridization assay can not be used. Besides a high throughput DNA sequencer and DNP chips, various methods and instruments have been developed for SNP analysis. They include the probe array assays with color-coded beads and bioluminometric assays such as Pyrosequencing and BAMPER (Bioluminometric Assay with Modified Primer Extension Reaction). It is demonstrated that BAMPER is very promising for realizing an inexpensive, cost effective and easy to use SNP typing method from the viewpoint of clinical diagnostics.View full abstractDownload PDF (5373K) -
Tsuneo Okuyama2001 Volume 45 Issue 4 Pages 227-230
Published: December 15, 2001
Released on J-STAGE: March 31, 2009
JOURNAL FREE ACCESSAn overview on the biotechnology and also analytical frontiers are summarized. In the analysis space defined, the limits of analysis concerning to the sensitivity and the separation were discussed. From this kind of discussions, we can develop the various ideas on the instrumentation and method. For bioentrepleneur we need various standards domestically and internationally. Under these considerations, a concept for the future image was presented on the activities in biotechnology.View full abstractDownload PDF (570K) -
2001 Volume 45 Issue 4 Pages 231-251
Published: December 15, 2001
Released on J-STAGE: March 31, 2009
JOURNAL FREE ACCESSDownload PDF (23818K) -
2001 Volume 45 Issue 4 Pages 253-281
Published: December 15, 2001
Released on J-STAGE: March 31, 2009
JOURNAL FREE ACCESSDownload PDF (23755K) -
Tadashi Hoshino, Tateshi Ojima, Toshihiro Nakamura, Kazunari Kumasaka, ...2001 Volume 45 Issue 4 Pages 283-286
Published: December 15, 2001
Released on J-STAGE: March 31, 2009
JOURNAL FREE ACCESSFour types of the hereditary variant human amylase (AMY) of pancreatic (P) origin have been reported as slow-P, dominant-P2, dominant-P2S, and dominant-P2MS by AMY isoenzyme on cellulose acetate membrane electrophoresis. Since 1990, we examined 84, 084 samples for serum and urine AMY isoenzymes, and found AMY bands which showed slower migration than P1-AMY, the main band of P-AMY, in 24 serum samples. These bands did not react with the anti-human salivary (S)-type AMY activity inhibitory monoclonal antibody, suggesting that these bands seemed to be derived from P-type AMY. When slow-P AMY bands were classified by the relative distance ratio of slow-P AMY and P1-AMY bands to P1-AMY and S1-AMY bands, it was found that there were 3 variants. The slow-P AMY variants were for convenience' sake named slow-P1 AMY (mean±1SD of the relative distance ratio: 0.518±0.043), slow-P2 AMY (0.635±0.024), and slow-P3 AMY (0.985±0.022) progressively from the anode side. The frequency of 3 slow-P AMY variants in Japanese is estimated at 0.007% (6/84, 084) for slow-P1 AMY, 0.019% (16/84, 084) for slow-P2 AMY, and 0.002% (2/84, 084) for slow-P3 AMY. At present, however, there is no evidence that these bands are genetic variants because pancreatic juice from patients and their family lines have not yet been examined.View full abstractDownload PDF (2145K) -
Shugo Nawata, Yoshinori Suminami, Hiroshi Hirakawa, Kenji Umayahara, H ...2001 Volume 45 Issue 4 Pages 287-289
Published: December 15, 2001
Released on J-STAGE: March 31, 2009
JOURNAL FREE ACCESSWe examined the heat-stability of tumor-associated serine protease inhibitor (serpin), squamous cell carcinoma (SCC) antigen by electrophoretic methods. SDS-PAGE and two-dimensional electrophoresis (2-DE) showed that SCC antigen-1 is a representative molecule of the heat stable cytosolic proteins in the tumor tissue extract. This unique heat stable property might be useful for the partial purification of SCC antigen-1.View full abstractDownload PDF (3424K)
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