Objective To investigate a 272 base pair section of the 5' non coding region of genomic DNA from the peripheral blood monounuclear cells of healthy hepatitis virus C (HCV) negative human subjects (not patien...Objective To investigate a 272 base pair section of the 5' non coding region of genomic DNA from the peripheral blood monounuclear cells of healthy hepatitis virus C (HCV) negative human subjects (not patients) Mothods This sequence section bears interest because ① it harbors several potential methylation (Cp rich) sites, and ② it represents the largest part of its internal ribosomal entry site A pre PCR digestion protocol was established making consistent use of four restriction endonucleases selected for certain features: SmaI, XmaCI, MspI, and HpaII are inhibited if methylation(s) are present at certain cytosines within their cutting sequences Results The suspected HCV specific sequence was found in the DNA of each subject tested The pre PCR digestion assay reveals individual differences in their pattern of methylation, which may be due to possible epigenetic phenomena Conclusions The results provide formal proof that these HCV specific sequences are contained in the genomic or extra chromosomal target DNA, and probably belong to a new class of endogenous sequences展开更多
The next generation sequencing enables generation of high resolution and high throughput data for structure sequence of any genome at a fast declining cost. This opens opportunity for population based genetic and geno...The next generation sequencing enables generation of high resolution and high throughput data for structure sequence of any genome at a fast declining cost. This opens opportunity for population based genetic and genomic analyses. In many applications, whole genome sequencing or re-sequencing is unnecessary or prohibited by budget limits. The Reduced Representation Genome Sequencing （RRGS）, which sequences only a small proportion of the genome of interest, has been proposed to deal with the situations. Several forms of RRGS are proposed and implemented in the literature. When applied to plant or crop species, the current RRGS protocols shared a key drawback that a significantly high proportion （up to 60%） of sequence reads to be generated may be of non-genomic origin but attributed to chloroplast DNA or rRNA genes, leaving an exceptional low efficiency of the sequencing experiment. We recommended and discussed here the design of optimized simplified genomic DNA and bisulfite sequencing strategies, which may greatly improves efficiency of the sequencing experiments by bringing down the presentation of the undesirable sequencing reads to less than 10% in the whole sequence reads. The optimized RAD- seq and RRBS-seq methods are potentially useful for sequence variant screening and genotyping in large plant/crop populations.展开更多
文摘Objective To investigate a 272 base pair section of the 5' non coding region of genomic DNA from the peripheral blood monounuclear cells of healthy hepatitis virus C (HCV) negative human subjects (not patients) Mothods This sequence section bears interest because ① it harbors several potential methylation (Cp rich) sites, and ② it represents the largest part of its internal ribosomal entry site A pre PCR digestion protocol was established making consistent use of four restriction endonucleases selected for certain features: SmaI, XmaCI, MspI, and HpaII are inhibited if methylation(s) are present at certain cytosines within their cutting sequences Results The suspected HCV specific sequence was found in the DNA of each subject tested The pre PCR digestion assay reveals individual differences in their pattern of methylation, which may be due to possible epigenetic phenomena Conclusions The results provide formal proof that these HCV specific sequences are contained in the genomic or extra chromosomal target DNA, and probably belong to a new class of endogenous sequences
文摘The next generation sequencing enables generation of high resolution and high throughput data for structure sequence of any genome at a fast declining cost. This opens opportunity for population based genetic and genomic analyses. In many applications, whole genome sequencing or re-sequencing is unnecessary or prohibited by budget limits. The Reduced Representation Genome Sequencing （RRGS）, which sequences only a small proportion of the genome of interest, has been proposed to deal with the situations. Several forms of RRGS are proposed and implemented in the literature. When applied to plant or crop species, the current RRGS protocols shared a key drawback that a significantly high proportion （up to 60%） of sequence reads to be generated may be of non-genomic origin but attributed to chloroplast DNA or rRNA genes, leaving an exceptional low efficiency of the sequencing experiment. We recommended and discussed here the design of optimized simplified genomic DNA and bisulfite sequencing strategies, which may greatly improves efficiency of the sequencing experiments by bringing down the presentation of the undesirable sequencing reads to less than 10% in the whole sequence reads. The optimized RAD- seq and RRBS-seq methods are potentially useful for sequence variant screening and genotyping in large plant/crop populations.
文摘应用限制性片段长度多态性（RFLP）、双向逆转录-聚合酶链式反应（BD-PCR）、序列分析等技术对我国部分地区柑橘衰退病毒（Citrus tristeza virus，CTV）柚类分离株进行了鉴定分析。结果表明：柚类以受相对单一的CTV株系侵染为主；柚类上的优势流行株系属于p25／Hin f ⅠRFLP6组群（占79．4％）和p23／BD-PCRⅢ组群（占84．1％）；在对柚类CTV分离株S051～S058及国外CTV分离株PB61、T30、T36、T385、SY568、VT、NUAGA的p23、p25基因序列分析中，S051与弱毒株PB61的同源性最高；S052与弱毒株T30、T385的同源性最高；S053、S054、S055、S056、S057、S058与其它CTV分离株的同源关系均较远，并在系统发生树上形成了独立的分枝。
