Though Yunnan province contains some 562 known species of fish,no cell lines from any of these have been made available to date.To protect germplasm resources and provide an effective tool in solving problems at cellu...Though Yunnan province contains some 562 known species of fish,no cell lines from any of these have been made available to date.To protect germplasm resources and provide an effective tool in solving problems at cellular level of Anabarilius grahami,a fish endemic to Fuxian Lake,Yunnan,China,we established and characterized the major features of a continuous cell line(AGF II)from the caudal fin tissue of A.grahami.This AGF II cell line consists of fibroblast-like cells and has been subcultured more than 60 times over the course of a year.The cell line was maintained in DMEM/F12 supplemented with 10%FBS,with a cellular doubling time of 51.1 h.We continued with more experiments to optimize the culture and storage conditions,and found a variety of interesting results:cells could grow at temperature between 24℃and 28℃,with the optimal temperature of 28℃.Likewise,the growth rate of A.grahami fin cells increased when the FBS proportion increased from 5%to 20%,with the optimal growth at the concentrations of 20%FBS;cells were able to grow in L-15 and DMEM/F12 with optimal growth at L-15;DMSO is a better cryoprotectant than Glycerol,EG and MeOH for AGFII cells with optimal concentration of 5%DMSO.Chromosome analysis also showed that the distribution of chromosome number varies from 38 to 52,with a modal peak at 48 chromosomes,accounting for 39.8%of all cells.Using the same primer pairs specific to mtDNA,the AGF II cell sequences obtained by PCR were identical to those from muscle tissues of A.grahami.Both chromosome analysis and PCR amplification confirmed the AGF II cells were from A.grahami,also indicating that that current long-term artificial propagation of A.grahami has been successful.Finally,we noted that when cells were transfected with pEYFP-N1 and pECFP-N1 plasmid,bright fluorescent signals were observed,suggesting that this cell line may be suitable for use in transfection and future gene expression studies。展开更多
Though Yurman province contains some 562 known species of fish, no cell lines from any of these have been made available to date. To protect germplasm resources and provide an effective tool in solving problems at cel...Though Yurman province contains some 562 known species of fish, no cell lines from any of these have been made available to date. To protect germplasm resources and provide an effective tool in solving problems at cellular level of Anabarilius grahami, a fish endemic to Fuxian Lake, Yunnan, China, we established and characterized the major features of a continuous cell line （AGF II） from the caudal fin tissue of A. grahami. This AGF II cell line consists of fibroblast-like cells and has been subeultured more than 60 times over the course of a year. The cell line was maintained in DMEM/F12 supplemented with 10% FBS, with a cellular doubling time of 51.1 h. We continued with more experiments to optimize the culture and storage conditions, and found a variety of interesting results： cells could grow at temperature between 24 ~C and 28 ~C, with the optimal temperature of 28 ~C. Likewise, the growth rate ofA. grahami fin cells increased when the FBS proportion increased from 5% to 20%, with the optimal growth at the concentrations of 20% FBS, cells were able to grow in L-15 and DMEM/FI2 with optimal growth at L-15; DMSO is a better eryoprotectant than Glycerol, EG and MeOH for AGFII cells with optimal concentration of 5% DMSO. Chromosome analysis also showed that the distribution of chromosome number varies from 38 to 52, with a modal peak at 48 chromosomes, accounting for 39.8% of all cells. Using the same primer pairs specific to mtDNA, the AGF II cell sequences obtained by PCR were identical to those from muscle tissues ofA. grahami. Both chromosome analysis and PCR amplification confirmed the AGF II cells were from A. grahami, also indicating that that current long-term artificial propagation ofA. grahami has been successful. Finally, we noted that when cells were transfected with pEYFP-N1 and pECFP-N1 plasmid, bright fluorescent signals were observed, suggesting that this cell line may be suitable for use in transfection and future gene expression studies.展开更多
基金the Global Environment Foundation/The World Bank Project(GEF-MSP grant No.TF051795)the Yunnan Development and Reform Commission。
文摘Though Yunnan province contains some 562 known species of fish,no cell lines from any of these have been made available to date.To protect germplasm resources and provide an effective tool in solving problems at cellular level of Anabarilius grahami,a fish endemic to Fuxian Lake,Yunnan,China,we established and characterized the major features of a continuous cell line(AGF II)from the caudal fin tissue of A.grahami.This AGF II cell line consists of fibroblast-like cells and has been subcultured more than 60 times over the course of a year.The cell line was maintained in DMEM/F12 supplemented with 10%FBS,with a cellular doubling time of 51.1 h.We continued with more experiments to optimize the culture and storage conditions,and found a variety of interesting results:cells could grow at temperature between 24℃and 28℃,with the optimal temperature of 28℃.Likewise,the growth rate of A.grahami fin cells increased when the FBS proportion increased from 5%to 20%,with the optimal growth at the concentrations of 20%FBS;cells were able to grow in L-15 and DMEM/F12 with optimal growth at L-15;DMSO is a better cryoprotectant than Glycerol,EG and MeOH for AGFII cells with optimal concentration of 5%DMSO.Chromosome analysis also showed that the distribution of chromosome number varies from 38 to 52,with a modal peak at 48 chromosomes,accounting for 39.8%of all cells.Using the same primer pairs specific to mtDNA,the AGF II cell sequences obtained by PCR were identical to those from muscle tissues of A.grahami.Both chromosome analysis and PCR amplification confirmed the AGF II cells were from A.grahami,also indicating that that current long-term artificial propagation of A.grahami has been successful.Finally,we noted that when cells were transfected with pEYFP-N1 and pECFP-N1 plasmid,bright fluorescent signals were observed,suggesting that this cell line may be suitable for use in transfection and future gene expression studies。
文摘Though Yurman province contains some 562 known species of fish, no cell lines from any of these have been made available to date. To protect germplasm resources and provide an effective tool in solving problems at cellular level of Anabarilius grahami, a fish endemic to Fuxian Lake, Yunnan, China, we established and characterized the major features of a continuous cell line （AGF II） from the caudal fin tissue of A. grahami. This AGF II cell line consists of fibroblast-like cells and has been subeultured more than 60 times over the course of a year. The cell line was maintained in DMEM/F12 supplemented with 10% FBS, with a cellular doubling time of 51.1 h. We continued with more experiments to optimize the culture and storage conditions, and found a variety of interesting results： cells could grow at temperature between 24 ~C and 28 ~C, with the optimal temperature of 28 ~C. Likewise, the growth rate ofA. grahami fin cells increased when the FBS proportion increased from 5% to 20%, with the optimal growth at the concentrations of 20% FBS, cells were able to grow in L-15 and DMEM/FI2 with optimal growth at L-15; DMSO is a better eryoprotectant than Glycerol, EG and MeOH for AGFII cells with optimal concentration of 5% DMSO. Chromosome analysis also showed that the distribution of chromosome number varies from 38 to 52, with a modal peak at 48 chromosomes, accounting for 39.8% of all cells. Using the same primer pairs specific to mtDNA, the AGF II cell sequences obtained by PCR were identical to those from muscle tissues ofA. grahami. Both chromosome analysis and PCR amplification confirmed the AGF II cells were from A. grahami, also indicating that that current long-term artificial propagation ofA. grahami has been successful. Finally, we noted that when cells were transfected with pEYFP-N1 and pECFP-N1 plasmid, bright fluorescent signals were observed, suggesting that this cell line may be suitable for use in transfection and future gene expression studies.
