In order to elucidate the molecular mechanisms of the oyster (Crassostrea ariakensis) against adverse stimulating factors, we cloned and sequenced a partial cDNA encoding a 70 kDa heat shock cognate protein (Hsc70) fr...In order to elucidate the molecular mechanisms of the oyster (Crassostrea ariakensis) against adverse stimulating factors, we cloned and sequenced a partial cDNA encoding a 70 kDa heat shock cognate protein (Hsc70) from the oyster. The live oysters were obtained from Chengcun, Yangxi County, Guangdong Province, China. Various tissues, including mantle, gills, adductor muscle, heart and blood cells, were respectively collected from 5 untreated live oysters or treated ones at 36℃ for 1 5 hours, and immediately frozen in liquid nitrogen except for the blood cells which were suspended with Trizol Reagent after centrifugation ( 12 000 r/min for 30 s) and stored at -20℃. Total RNA was isolated using Trizol Reagent according to the manufacture’s instructions. The first strand cDNA was synthesized using reverse transcriptase Superscript Ⅱ according to the manufacture’s instructions. The primers were designed from a conserved region of C. gigas Hsc70 cDNA sequence (GeneBank accession No. AF144646). The polymerase chain reaction (PCR) was performed for 30 cycles with denaturation at 94℃ for 30 s, annealing at 49℃ for 40 s, and elongation at 72℃ for 30 s. The product was cloned to pGEM T easy vector and sequenced. It is 509 base pairs (bp) and possesses 94% identity with the cDNA encoding C. gigas Hsc70 using Blastn. This homology was strongly confirmed by amino acid sequence comparison using the Blastx (99%). The 509 bp fragment was labeled with α 32 pdCTP and a random primer DNA labeling kit and employed as a probe to perform Southern blotting, the result demonstrated that the cDNA came from a partial mRNA transcript of C. ariakensis genomic DNA gene. The polymerase chain reaction (PCR) was carried out to investigate the expression of Hsc70, Using the cDNAs of several tissues, such as gills (heat shocked), mantle, adductor muscle (heat shocked), heart, blood cells (one sample with heat shock for 1 5 hours at 36℃ and another without any stimulus). The PCR results revealed that Hsc70 transcripts could be detected in all the tissues analyzed and greatly increased in the tissues with heat shock. The results showed that the Hsc70 is ubiquitously and constitutively expressed but can be stimulated by heat shock. All the facts above firmly established that the cloned cDNA fragment was a part of the cDNA encoding a Hsc70 protein in the oyster C. ariakensis .展开更多
为了揭示2001-2014年间Southern印迹杂交领域的研究现状及发展趋势,文章以Web of ScienceTM核心合集数据库为数据源,利用CiteSpaceⅢ对Southern印迹杂交领域相关论文进行了共被引分析.结果发现:该领域研究热点以基础应用为主,核心作者...为了揭示2001-2014年间Southern印迹杂交领域的研究现状及发展趋势,文章以Web of ScienceTM核心合集数据库为数据源,利用CiteSpaceⅢ对Southern印迹杂交领域相关论文进行了共被引分析.结果发现:该领域研究热点以基础应用为主,核心作者群明显,研究力量缺乏合作,需寻找新的研究热点,促进该领域发展.展开更多
从东乡野生稻(O ry za ruf ipogon G riff)中克隆到7个重复序列,序列分析表明:这些序列是与转座因子有关的序列。以序列H 2为探针,对不同基因组水稻进行Southern杂交,表明该序列为稻属内AA基因组特异的重复序列。其分布在水稻的多条染...从东乡野生稻(O ry za ruf ipogon G riff)中克隆到7个重复序列,序列分析表明:这些序列是与转座因子有关的序列。以序列H 2为探针,对不同基因组水稻进行Southern杂交,表明该序列为稻属内AA基因组特异的重复序列。其分布在水稻的多条染色体上,荧光原位杂交(F ISH)结果显示该重复序列主要位于染色体的着丝粒以及端粒附近,重复单位序列长度为615 bp左右。并就特异重复序列在水稻研究中的应用进行了讨论。展开更多
文摘In order to elucidate the molecular mechanisms of the oyster (Crassostrea ariakensis) against adverse stimulating factors, we cloned and sequenced a partial cDNA encoding a 70 kDa heat shock cognate protein (Hsc70) from the oyster. The live oysters were obtained from Chengcun, Yangxi County, Guangdong Province, China. Various tissues, including mantle, gills, adductor muscle, heart and blood cells, were respectively collected from 5 untreated live oysters or treated ones at 36℃ for 1 5 hours, and immediately frozen in liquid nitrogen except for the blood cells which were suspended with Trizol Reagent after centrifugation ( 12 000 r/min for 30 s) and stored at -20℃. Total RNA was isolated using Trizol Reagent according to the manufacture’s instructions. The first strand cDNA was synthesized using reverse transcriptase Superscript Ⅱ according to the manufacture’s instructions. The primers were designed from a conserved region of C. gigas Hsc70 cDNA sequence (GeneBank accession No. AF144646). The polymerase chain reaction (PCR) was performed for 30 cycles with denaturation at 94℃ for 30 s, annealing at 49℃ for 40 s, and elongation at 72℃ for 30 s. The product was cloned to pGEM T easy vector and sequenced. It is 509 base pairs (bp) and possesses 94% identity with the cDNA encoding C. gigas Hsc70 using Blastn. This homology was strongly confirmed by amino acid sequence comparison using the Blastx (99%). The 509 bp fragment was labeled with α 32 pdCTP and a random primer DNA labeling kit and employed as a probe to perform Southern blotting, the result demonstrated that the cDNA came from a partial mRNA transcript of C. ariakensis genomic DNA gene. The polymerase chain reaction (PCR) was carried out to investigate the expression of Hsc70, Using the cDNAs of several tissues, such as gills (heat shocked), mantle, adductor muscle (heat shocked), heart, blood cells (one sample with heat shock for 1 5 hours at 36℃ and another without any stimulus). The PCR results revealed that Hsc70 transcripts could be detected in all the tissues analyzed and greatly increased in the tissues with heat shock. The results showed that the Hsc70 is ubiquitously and constitutively expressed but can be stimulated by heat shock. All the facts above firmly established that the cloned cDNA fragment was a part of the cDNA encoding a Hsc70 protein in the oyster C. ariakensis .
文摘为了揭示2001-2014年间Southern印迹杂交领域的研究现状及发展趋势,文章以Web of ScienceTM核心合集数据库为数据源,利用CiteSpaceⅢ对Southern印迹杂交领域相关论文进行了共被引分析.结果发现:该领域研究热点以基础应用为主,核心作者群明显,研究力量缺乏合作,需寻找新的研究热点,促进该领域发展.
文摘从东乡野生稻(O ry za ruf ipogon G riff)中克隆到7个重复序列,序列分析表明:这些序列是与转座因子有关的序列。以序列H 2为探针,对不同基因组水稻进行Southern杂交,表明该序列为稻属内AA基因组特异的重复序列。其分布在水稻的多条染色体上,荧光原位杂交(F ISH)结果显示该重复序列主要位于染色体的着丝粒以及端粒附近,重复单位序列长度为615 bp左右。并就特异重复序列在水稻研究中的应用进行了讨论。
