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S T Liu

Publications and source records attributed to S T Liu.

At least 55 records · Page 3Linked to original sources
PubMed1995

Transposon mutagenesis and cloning of the genes encoding the enzymes of fengycin biosynthesis in Bacillus subtilis.

A total of 20 Bacillus subtilis F29-3 mutants defective in fengycin biosynthesis was obtained by Tn917 mutagenesis. Cloning and mapping results showed that the transposon in these mutants was inserted in eleven different locations on the chromosome. We were able to use the chromosomal sequence adjacent to the transposon as a probe to screen for cosmid clones containing the fengycin biosynthesis genes. One of the clones obtained, pFC660, was 46 kb long. Eight transposon insertion sites were mapped within this plasmid. Among the eleven different mutants analyzed, four mutants had Tn917 inserted in regions which encoded peptide sequences similar to part of gramicidin S synthetase, surfactin synthetase, and tyrocidine synthetase. Our results suggest that fengycin is synthesized nonribosomally by the multienzyme thiotemplate mechanism.

Adenosine Triphosphatases↗
PubMed1995

Sequence analysis of an Erwinia stewartii plasmid, pSW100.

The nucleotide sequence of the smallest plasmid of Erwinia stewartii SW2 was determined. This plasmid, pSW100 (4272 bp), consists of a 702-bp region homologous to the origins of replication of plasmids p15A, ColE1, and ColA. Plasmid pSW100 also contains sequences homologous to the bom region and mobCABD genes of ColE1, except that a single-base deletion in mobA has been detected. This deletion did not affect the mobilization ability of pSW100 by an endogenous conjugative plasmid of E. stewartii SW2, pDC250. Plasmid pSW100 also has a 1596-bp open reading frame with five 132-bp perfect repeats. However, this open reading frame and the repeats are not required for plasmid replication. Furthermore, curing of pSW100 did not cause any phenotypic change, suggesting that this plasmid is not essential for the survival of the organism.

Amino Acid Sequence↗
PubMed1995

Characterization of 5'-upstream sequence of the latent membrane protein 1 (LMP-1) gene of an Epstein-Barr virus identified in nasopharyngeal carcinoma tissues.

Sequence variations of the 5'-upstream region of latent membrane protein 1 (LMP-1) in two Epstein-Barr virus (EBV) strains have been reported before (Chen et al., 1992). To investigate the effect of these variations on gene expression, we constructed a series of deletion plasmids encompassing positions -950 to +20 of the LMP-1 promoter region and tested for the ability to drive chloramphenicol acetyltransferase (CAT) gene expression in C33A cells. Results showed that the promoter activities of constructs from NPC strain were 3-fold lower than the corresponding constructs from the B95-8 strain. In addition, the region between -54 and +20 contained the basic, constitutive promoter activity for both strains. Sequence analysis of this region indicated that an activating transcription factor (ATF) binding site, TGACGTAG, which is present in B95-8 strain was changed to TCTCGTAG in NPC strain. A chimeric plasmid study suggested that these sequence variations in the ATF binding site may contribute to the 3-fold increase of CAT activity observed for B95-8 strain. Furthermore, the activity of the promoter constructs was not activated by EBV-encoded nuclear antigen 2 (EBNA-2) in C33A cells. However, the promoter activities were upregulated in B-lymphocyte cells such as CG3 and CA46 cells. The biological significance of this difference in promoter activity of LMP-1 gene between two strains and the involvement of the cellular factors were discussed.

Base Sequence↗
PubMed1995

Identification of an internal promoter of the latent membrane protein 1 gene of Epstein-Barr virus.

The latent membrane protein 1 (LMP 1) gene of an Epstein-Barr virus (EBV) variant derived from an nasopharyngeal carcinoma (NPC) biopsy in Taiwan was isolated and characterized (Chen et al., 1992). Transient expression of the genomic sequence containing this gene showed two proteins with molecular masses of 62 kD and 50 kD, respectively, recognized by LMP-specific antibody S12. To determine if these two proteins were derived from independent promoters, we generated a series of mutant plasmids from plasmid pT7(E) that contained the upstream and the coding regions of the LMP 1 gene. These mutants were introduced into a human epithelial cell line, C33A, and LMP 1 proteins were examined by Western blotting analysis with the S12 antibody. Data showed that plasmid with a fragment containing approximately 3 kb of upstream sequence of LMP 1 gene produced the 62-kD protein. Removal of 2.7 kb of the upstream sequence (plasmid delta 2710, deleted to nucleotide 169,571) resulted in the production of both 62-kD and 50-kD proteins. This suggested that the upstream sequence interfered with the production of the 50-kD protein. Plasmid DNA deleted to Acc I site (nucleotide 169,223) generated only the 50-kD protein, designated as tr-LMP. Further deletion to nucleotide 169,038 resulted in the expression of another smaller LMP1 (49 kD, named as str-LMP1). The region between nucleotides 168,789 and 169,038 was tested to see if it possessed a promoter activity by using the luciferase gene as a reporter. Data showed that this region contained promoter activity with a level compatible to the previously reported ED-L1A promoter.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1995

Identification of a novel promoter located within the Bam HI Q region of the Epstein-Barr virus genome for the EBNA 1 gene.

EBNA 1 is the only antigen expressed in both Epstein-Barr virus (EBV)-infected nasopharyngeal carcinoma (NPC) and Burkitt's lymphoma cells. Previous studies showed that the mRNA of EBNA 1 in these two tumor tissues was initiated from a promoter located in the Bam HI F fragment (Fp) on the viral genome. Two regulatory elements located in the downstream Bam HI Q region include an EBNA 1 binding site and a positive regulatory region between the Fp and the EBNA1 binding site. This data strongly suggested that a cellular factor(s) may modulate the usage of the Fp. To locate the shortest responsible viral sequence, we constructed a series of luciferase gene and chloramphenicol acetyltransferase (CAT) gene plasmids that contained various portions of the Bam HI F/Q region. Plasmid DNA was then introduced into cells to examine the promoter activity of each construct. By this method, we identified a 186-bp fragment within the Bam HI Q region that possessed the highest activity. This promoter was designated as Qp and found to be orientation-dependent and down-regulated by EBNA 1 in both the type I BL cells and human epithelial cells. Furthermore, RNase protection assay showed that a transcription initiation site was located at nucleotide 62,416 of the EBV genome. Reverse transcriptase polymerase chain reaction (RT-PCR) analysis further confirmed that the transcript was initiated from the Qp, and not the Fp. Therefore, our data suggested that a novel promoter, Qp, located within the Bam HI Q existed for the EBNA 1 expression in the latently infected type 1 BL cells. The biological significance of the selection of the Qp needs further investigation.

Antigens, Viral↗
PubMed1994

Construction of Tn917ac1, a transposon useful for mutagenesis and cloning of Bacillus subtilis genes.

A Tn917 derivative was constructed for the purposes of mutagenesis and cloning of Bacillus subtilis genes. This transposon, Tn917ac1 (4.6 kb), consisted of terminal inverted repeats of Tn917, the res sequence, a ColE1 origin of replication (ori) and two drug-resistance genes. The plasmid carrying this transposon, named pD917, contained the erm-tnpR-tnpA gene cluster of Tn917 and a temperature-sensitive ori of pE194. For the purpose of mutagenesis, transposition of Tn917ac1 was induced by culturing strains harboring pD917 in a medium containing a low concentration of erythromycin. Cells with a Tn917ac1 insertion in the chromosome were selected on agar containing chloramphenicol after heat treatment to eliminate the plasmidic form of pD917. DNA fragments adjacent to Tn917ac1 could be cloned by restriction digestion of the chromosomal DNA and by transforming the self-ligated restriction fragments into Escherichia coli. Sequence analysis revealed that Tn917ac1 was integrated into the chromosome of B. subtilis by transposition in a recE strain and by transposition or integration of pD917 in a wild-type strain. Tn917ac1 has been demonstrated to be useful for mutating and cloning of the genes involved in the biosynthesis of fengycin in B. subtilis F29-3.

Bacillus subtilis↗
PubMed1994

Transcriptional activation of flanking sequences by Tn1000 insertion.

The carotenoid biosynthesis operon of Erwinia herbicola Eho13 consists of five genes, which are organized in the order crtE-crtX-crtY-crtI-crtB. These genes, with the exception of crtX, encode functions of beta-carotene biosynthesis and give an orange-coloured phenotype in Escherichia coli. Since crtX is not involved in the biosynthesis of beta-carotene, deletion of this gene does not alter the phenotype of pigmented cells. On the other hand, insertion of Tn1000 into crtX or into the upstream untranslated region of the operon resulted in a light-yellow, rather than an unpigmented phenotype, indicating that Tn1000 does not exert a strong polar effect when inserted in this operon. RNA analysis revealed that the sequence downstream from the insertion site was transcribed at a low level. Primer extension showed that the "-35"-like sequence in the terminal inverted repeats was not responsible for the transcription of the downstream sequence. Furthermore, primer extension and polymerase chain reaction (PCR) studies revealed that RNA transcribed from the promoters inside of Tn1000 was extended through the terminal inverted repeats into the adjacent sequences. In addition Tn1000, in either orientation, was able to generate fusion transcripts when placed upstream of a promoter-less tetracycline-resistance gene and resulted in cells resistant to the drug. These results showed that Tn1000 insertion transcriptionally activates the DNA sequences adjacent to the transposon.

Base Sequence↗
PubMed1994

Analysis of the gene cluster encoding carotenoid biosynthesis in Erwinia herbicola Eho13.

Erwinia herbicola is known to synthesize carotenoids and gives an orange-coloured phenotype. These carotenoids play a role in the protection of the cells from the damage caused by near-UV irradiation in nature. The genes encoding these carotenoids in E. herbicola Eho13 are clustered in a 7 kb DNA fragment. The complete sequence of this fragment has been determined. DNA sequence analysis revealed that the entire sequence contains at least five genes, which are transcribed in the same direction. These five genes are organized in the order crtE-crtX-crtY-crtI-crtB. A gene fusion study showed that two different regions in this 7 kb gene cluster contain promoter activity. Primer-extension analysis identified two transcription start sites, located 147 bp upstream from the first gene of the cluster, crtE, and within the last gene of the cluster, crtB. An RNA-PCR study suggested that the five crt genes were organized in an operon and were transcribed from the promoter upstream from crtE.

Alkyl and Aryl Transferases↗
PubMed1994

Prevalence of Taiwan variant of Epstein-Barr virus in throat washings from patients with head and neck tumors in Taiwan.

The prevalence of the Epstein-Barr virus (EBV) Taiwan variant was investigated in the throat washing (TW) samples from patients with head and neck tumors, persons with nonmalignant diseases, and healthy adults in Taiwan. By using the EBV (BNLF-1 gene)-specific primers and PCR, the EBV latent membrane protein gene BNLF-1 was detected in 91 (61%) of the 150 TW samples from patients with tumors, including 25 (78%) of 32 patients with nasopharyngeal carcinoma and 66 (56%) of 118 other patients with head and neck tumors. The TW samples from the 26 patients with nonmalignant tumors and 53 healthy adults were also examined. Approximately 47% of these samples were positive for the EBV gene. The PCR products of the BNLF-1 gene were then subjected to XhoI digestion. Sixty-eight of 91 PCR products (75%) showed the loss of the XhoI site, which indicated the presence of a Taiwan strain of EBV in patients with tumors. The DNA sequence of the BNLF-1 gene of the Taiwan variant revealed that the loss of the XhoI site was due to a nucleotide change from a G to a T at position 169,426 in comparison with the sequence of prototype EBV B95-8 cells. Furthermore, the Taiwan strain appeared significantly more frequently in the TWs and tissue samples from patients with nasopharyngeal carcinoma (88%; P < 0.001) and laryngeal carcinoma (80%; P < 0.02) than in those samples from healthy adults (about 40%). These data indicate that a Taiwan variant of EBV may be closely associated with head and neck tumors and suggest that this variant may be important in the pathogenesis of head and neck tumors.

Base Sequence↗
PubMed1994

Congenital cutaneous candidiasis--report of two cases.

Candida infection contracted in utero and manifested at birth is a rare event. Two newborn infants are presented with generalized maculopapular rashes at birth. The eruption soon became vesicles or pustules; no other manifestations were found. Candida albicans was isolated from the skin lesion. After topical antifungal application, the lesions became fine and desquamated in the recuperative period. No complication was noted during the course. The diagnosis of congenital cutaneous candidiasis relies on its clinical course and positive culture result of the skin lesion. No systemic antifungal therapy is recommended except for the high risk infants.

Candidiasis, Cutaneous↗
PubMed1994

Light-emitting aeromonas and plesiomonas generated by transconjugating luxAB from Escherichia coli.

The transposon derivative has been placed on a transposition suicide vector to yield pDB30 in Escherichia coli WA803. A simple method, using a Tn5 derivative Tn5-Lux, has been successfully devised for the introduction and stable expression of the bioluminescence property in Pseudomonas sp., Agrobacterium sp., and Rhizobium sp. In this study, there was also successful mating between Escherichia coli WA803(pDB30) and strains of Acromonas hydrophila and Plesiomonas shigelloides. These bacteria emitted bioluminescence after they gained pDB30 by transconjugation.

Aeromonas↗
PubMed1993

Carotenoid-biosynthesis genes as a genetic marker for the purpose of gene cloning.

A cloning vector pSL775 (7.0 kb) was constructed using the carotenoid-biosynthesis genes of Erwinia herbicola Eho13 (ATCC 53489). This vector contained a ColE1 origin, an ampicillin resistant gene, and a total of 11 single cloning sites: Asp718, AvaI, BamHI, BanII, EcoRV, HindIII, KpnI, MluI, NcoI, NotI, and SmaI. Transforming the vector into an Escherichia coli strain could result in pink clones. On the other hand, insertion of a DNA fragment into one of these cloning sites resulted in nonpigmented clones. The color differential between the two types of colonies could be detected visually on agar medium after culturing the cells at 37 degrees C for 18 hours.

Base Sequence↗
PubMed1993

Detection of point mutations in the chloroplast genome by single-stranded conformation polymorphism analysis.

Single-stranded conformation polymorphism (SSCP) analysis was used to examine the mutations of the chloroplast 16S rRNA locus of streptomycin-resistant mutants in Nicotiana plumbaginifolia. DNA fragments of 121, 517, 968 and 1578 bp, each possessing a known point mutation, were generated by polymerase chain reaction (PCR). The resulting fragments were denatured and separated by nondenaturing polyacrylamide gel electrophoresis. Compared to the patterns of the wild-type DNA fragments, the bands of the single-stranded DNA fragments of 121 and 517 bp with base changes were shifted. However, no pattern variations were detected from the DNA fragments of 968 and 1578 bp generated from both wild-type and mutants.

Base Sequence↗
PubMed1993

Detection of Epstein-Barr virus and human papillomavirus in head and neck tumors.

The presence of Epstein-Barr virus (EBV) DNA and human papillomavirus (HPV) DNA in 74 head and neck tumor tissues was examined by the polymerase chain reaction and DNA sequencing analysis. EBV DNA sequence was detected in all 30 nasopharyngeal-carcinoma tissue samples and in 30 of 44 other head and neck tumor samples. HPV DNA sequence was detected in 14 of 30 nasopharyngeal-carcinoma tissue samples and in 11 of 44 other tumor samples. Coinfection of both viruses was observed in 14 nasopharyngeal-carcinoma tissue samples but only in 5 other head and neck tumor samples including 3 hypopharyngeal-carcinoma tissue samples. Our data indicate that EBV is closely associated with nasopharyngeal- carcinoma and may also be related to hypopharyngeal-carcinoma. In addition, a relatively high percentage of EBV-positive nasopharyngeal- and hypopharyngeal-carcinoma tissue specimens contained HPV sequence. The significance of the coexistence of EBV and HPV in these tumor tissues requires further study.

Base Sequence↗
PubMed1992

Detection of mutations in the p53 gene in human head and neck carcinomas by single strand conformation polymorphism analysis.

Using the polymerase chain reaction (PCR)-based single strand conformation polymorphism (SSCP) analysis, we have examined the highly conserved regions of the p53 gene in 58 biopsy samples of head and neck tumors. Mutations were found in 13/58 (23%) tumor specimens, but not in 6 normal tissues. Ten of 13 mutations were due to single base changes and the remaining 3 were 1- or 8-base deletion mutants. These mutations were clustered in exons 5 and 7 and resulted in amino acid changes. Our results seem to indicate that mutations in the p53 gene contribute to a significant number of cases of the head and neck tumors including 20% of nasopharyngeal carcinoma biopsies. The relationship of Epstein-Barr virus or human papillomavirus and p53 gene mutations in this group of cancers was also analyzed and discussed.

Base Sequence↗
PubMed1992

Direct sequencing of tobacco chloroplast genome by the polymerase chain reaction.

We have developed a polymerase chain reaction (PCR) method for sequencing of tobacco chloroplast genome. In a mixture containing chloroplast DNA, 5'-end-labeled oligonucleotide primer, Taq DNA polymerase and reaction buffer, we were able to sequence a segment of chloroplast 16S rRNA gene. The results showed that the 750 bp of DNA sequenced were identical to the sequence reported, indicating that direct sequencing method that we have developed is useful for the sequencing of chloroplast genome. To analyze the chloroplast genome more rapidly in those in vitro grown plantlets, we also developed a simple method which is applicable for the amplifications and sequencing of chloroplast 16S rRNA fragment from either 0.15 g of tobacco leaf or stem tissue. The readable sequences obtained from the presented methods were consistent with the published sequence.

Base Sequence↗
PubMed1992

Distribution of type A and type B EBV in normal individuals and patients with head and neck carcinomas in Taiwan.

The subtypes of Epstein-Barr virus (EBV) according to the EBNA 2 gene were investigated in Taiwan by the polymerase chain reaction (PCR) and by Southern blot hybridization. The materials included 53 nasopharyngeal carcinoma (NPC) biopsies, 49 other head and neck cancers and 32 throat washings of normal individuals. EBV DNA was found in all NPC biopsies, 27 of 49 other head and neck carcinomas and 81% of normal individuals. Type A EBV was the predominant type of EBV in both normal individuals and patients with head and neck carcinomas in Taiwan. Type B EBV or coexistence of the A and B types comprised a small number of samples in this study.

Base Sequence↗