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分子生物实验室常用的小公式

发布: 2017-04-26 1,382阅读 214评论

老师从小就教导我们：“先把数据算算对，再去做实验！还有，说的就是你啦，别忘了单位！”。不知道长做分子生物实验的同学们有没有这样的感觉，计算其实是一个十分重要的过程，这关系到整个实验的成功与否。有时候要算个摩尔数啊，再来个摩尔比啊，最后再来个单位换算什么的，一堆一堆，绝对不难。

切入正题，到底哪些公式会比较常用呢？小编认为有一下几种：

1. 摩尔浓度（mol/L）与溶质质量（g）

m (g) = Desired Molarity (mol/L) x V溶剂 (L) x M溶质 (g/mol)

2. 稀释倍数n 和稀释液的体积Vadding

n =（Cfinal * Vfinal）/（Cstock * Vstock）

Vadding = Vfinal – Vstock

Final=最终溶液

Stock=储备液/原液

3. 论OD260的重要性（当实验室的Nanodrop坏掉了或被坏人持续占用了）

CDNA/RNA (μg/ml) = Absorbance (OD260) * conversion factor
conversion factors:

  • 1 OD260 Unit = 50 μg/ml， dsDNA
  • 1 OD260 Unit = 40 μg/ml， ssRNA
  • 1 OD260 Unit = 33 μg/ml， ssDNA

这里的μg/ml也就是ng/μl，所以和Nanodrop测出来的单位是相同的。值得注意的是，这里的每一个OD260­单位对应的核酸浓度只是估算值，毕竟碱基组成不同也会造成折光度的差异，所以在有的网站上，这三组值也会略有差异。

4. DNA/RNA 质量到摩尔数的估测转换

dsDNA:

molesdsDNA (mol) = mdsDNA (g)/[(length of dsDNA (bp) x 617.96 g/mol) + 36.04 g/mol]

ssDNA:

molesssDNA (mol) = mssDNA (g)/[(length of ssDNA (nt) x 308.97 g/mol) + 18.02 g/mol]

ssRNA:

molesssRNA (mol) = mssRNA (g)/[(length of ssRNA (nt) x 321.47 g/mol) + 18.02 g/mol]

这里bp就是碱基对数，nt就是核苷酸数。

正常估算来讲一对碱基的平均摩尔质量是650 g/mol，小编这里用的是NEB （New England Biolad）推出的修正公式。617.96 g/mol 其实是除去了每对碱基两端的水分子（聚合成链的过程是要除去一个H和一个OH的）。最后又加上36.04 g/mol，是假设两个-OH和两个-H分别加在最后大分子的两端。这样的算法比直接使用650 g/mol要稍微准确一点，但是整个过程还是建立在估算碱基对平均分子量的基础之上，所以不论哪种方法计算出来的都是平均值而已。

5. 粘合反应（ligation）插入片段和质粒的摩尔比例

Massinsert (g) = (insert/vector molar ratio) x massvector (g) x (ratio of insert/vector lengths)

一般这个插入片段对质粒的摩尔比例可以是2:1或者是3:1，都可以根据自己的实验而定。另外一个重要的比例就是插入片段对质粒的序列长度比。

6. 单位换算

1 ng = 10-3 μg =10-6 mg = 10-9 g = 10-12 kg

1 cm3 = 10-3 dm3 = 10-6 m3

1 cm3 = 1 mL

1 dm3 = 1 L

等等等……

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