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- 详细信息
- 文献和实验
- 技术资料
- 库存:
60
- 英文名:
L-Carnitine inner salt
- CAS号:
541-15-1
- 供应商:
上海莼试
- 规格:
10mM (in 1mL DMSO) 20mg
| 产品名称 | L-Carnitine inner salt | 产品货号 | CS-01Y64255 |
| 规格 | 10mM (in 1mL DMSO) 20mg | CAS号 | 541-15-1 |
| 含量 | >98.00% | 分子式 | C7H15NO3 |
| 分子量 | 161.2 | 用途 | 仅供科研研究使用 |
L-carnitine (Levocarnitine) is constituent of striated muscle and liver. It is used therapeutically to stimulate gastric and pancreatic secretions and in the treatment of hyperlipoproteinemias.Target: OthersL-carnitine (Levocarnitine) is an endogenous molecule involved in fatty acid metabolism, biosynthesized within the human body using amino acids: L-lysine and L-methionine, as substrates. L-carnitine (Levocarnitine) can also be found in many foods, but red meats, such as beef and lamb, are the best choices for adding carnitine into the diet [1]. Administering L-carnitine (510 mg/day) to patients with the disease. L-carnitine (Levocarnitine) treatment significantly improved the total time for dozing off during the daytime, calculated from the sleep logs, compared with that of placebo-treated periods. L-carnitine (Levocarnitine) efficiently increased serum acylcarnitine levels, and reduced serum triglycerides concentration [2]. L-carnitine (Levocarnitine) and its derivatives show promise in the treatment of chronic conditions and diseases associated with mitochondrial dysfunction but further translational studies are needed to fully explore their potential [3].References:[1]. Marcovina, S.M., et al., Translating the basic knowledge of mitochondrial functions to metabolic therapy: role of L-carnitine. Transl Res, 2013. 161(2): p. 73-84.[2]. Pekala, J., et al., L-carnitine--metabolic functions and meaning in humans life. Curr Drug Metab, 2011. 12(7): p. 667-78.[3]. Miyagawa, T., et al., Effects of oral L-carnitine administration in narcolepsy patients: a randomized, double-blind, cross-over and placebo-controlled trial. PLoS One, 2013. 8(1): p. e53707.
化学性质:
规格:10mM (in 1mL DMSO) 20mg
CAS:541-15-1
别名:CARBOXY-N,N,N-TRIMETHYL-2-PROPEN-1-AMINIUM CHLORIDE;
化学名:(3R)-3-hydroxy-4-(trimethylazaniumyl)butanoate
分子式:C7H15NO3
分子量:161.2
溶解度:Ethanol: 10 mg/ml,PBS (pH 7.2): 10 mg/ml
储存条件:
General tips:For obtaining a higher solubility , please warm the tube at 37 ℃ and shake it in the ultrasonic bath for a while.
Shipping Condition: Evaluation sample solution : ship with blue ice All other available size: ship with RT , or blue ice upon request
使用方法:
1. 常用筛选浓度
注意:用来筛选稳转株的工作浓度需要根据细胞类型,培养基,生长条件和细胞代谢率而变化,推荐使用浓度为50-1000μg/mL。对于第一次使用的实验体系建议通过建立杀灭曲线(kill curve),即剂量反应性曲线,来确定最佳筛选浓度。
一般而言,哺乳动物细胞50-500μg/mL;细菌/植物细胞20-200μg/mL;真菌300-1000μg/mL。
2. 杀灭曲线的建立
注意:为了筛选得到稳定表达目的蛋白的细胞株,需要确定能够杀死未转染宿主细胞的抗生素浓度,可通过建立杀灭曲线(剂量反应曲线)来实现,至少选择5个浓度。
1) 第一天:未转化的细胞按照20-25%的细胞密度铺在合适的培养板上,37℃,CO2培养过夜;注:对于需要更高密度来检测活力的细胞,可增加接种量。
2) 根据细胞类型,设定合适范围内的浓度梯度。以哺乳动物细胞为例,可设定50,100,250,500,750,1000μg/mL。先用去离子水或者PBS buffer按照1:10的比例将母液稀释到5 mg/ml,然后按照下表稀释到相应浓度的工作液。
3) 第二天:替换旧的培养基,换用新鲜配制的含有相应浓度药物的培养基。每个浓度做三个平行孔。
4) 接下来每3-4天更换新的含药物培养基。
5) 按照固定的周期(如每2天)进行活细胞计数来确定阻止未转染细胞生长的恰当浓度。选择在理想的天数(通常是7-10天)内能够杀死绝大多数细胞的浓度为稳定转染细胞筛选用的工作浓度。
3. 稳定转染细胞的筛选
1) 转染48h后,用含有适当浓度的潮霉素B筛选培养基来传代细胞(直接传代或者稀释后传代)。
注意:细胞处于活跃分裂状态时抗生素的杀伤。则当细胞过于稠密,其效率会降低。为了得到较好的筛选效果,最好将细胞稀释至丰度不超过25%
2) 每隔3-4天更换含有药物的筛选培养液。
3) 筛选7天后观察并评估细胞克隆(集落)的形成情况。集落的形成可能还需要一周或者更多的时间,这取决于宿主细胞类型,转染,以及筛选效果。
4) 挑取并转移5-10个抗性克隆于35mm细胞培养板,继续用含药物的筛选培养液维持培养7天。
5) 之后更换正常培养基培养即可。
蛋白酶抑制剂混合物实验步骤:
(1)L-Carnitine inner salt实验开始前将RNA提取液于65℃水浴锅中预热,离心管中加入ME(巯基乙醇),(10mL加80ul,50mL中加入300ul)
(2)取约0.8g菌丝体(液体培养获得的菌丝用真空抽滤即可!固体培养就更好说了),在液氮中迅速磨成精细粉末,装入50mL离心管,按1g材料8mL的量加入预热的RNA提取液,颠倒混匀
(3)65℃水浴3-10 min,期间混匀2-3次
(4)加入等体积的酚(注意是酸酚pH4.5)::yi戊醇(25:24:1)抽提(10,000rpm,4℃,5 min)
(5)取上清,等体积的yi戊醇(24:1)抽提(10,000rpm,4℃,5 min)
(6)加入1/4V体积10M LiCl溶液,4℃放置6h以上(或过夜)
(7)10,000rpm,4℃离心20min
(8)弃上清,用500ul SSTE溶解沉淀
(9)酚::yi戊醇(25:24:1)抽提两次,:yi戊醇(24:1)抽提1次(10,000rpm,4℃,5min)
(10)加2V体积的无水乙醇,在-70℃冰箱沉淀30min以上
(11)12,000rpm,4℃离心20 min
(12)弃上清.沉淀用70%酒精漂洗一次,干燥
(13)加200ul的DEPC处理水溶解
(14)用非变性琼脂糖凝胶电泳和紫外分光光度计扫描检测RNA的质量(在抽提过程中,若蛋白质含量或其它的杂质还较多,可以增加抽提次数)
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