| T751065 |
Component |
200 U |
1000 U |
Storage |
| T751065A |
Thermostable RNase H |
200 U |
5×200 U |
-20°C. Avoid freeze/thaw cycle |
| T751065B |
10× RNase H Reaction Buffer |
1 mL |
5×1 mL |
-20°C. Avoid freeze/thaw cycle | |
储存缓冲液：500mM Tris-HCl, 750mM KCl, 100mM DTT, 30mM MgCl2, pH 8.3, 25°C
Thermostable RNase H，耐热 RNase H，是一种在较高温度(65°C以上)时仍然保持活性的核糖核酸内切酶(endoribonuclease)。Thermostable RNase H可以选择性识别并酶切 RNA:DNA 杂合双链中的 RNA 链，同时杂合双链中的 DNA 会保持完整。ThermostableRNase H 不会降解单链或双链的 RNA 或 DNA。
产品应用
1、高精度 RNA 结构图谱制备和 mRNA 加帽率分析。
2、cDNA 第二链合成中去除 mRNA;提高 RT-qPCR 效率。
3、在 Oligo(dT) 存在下除去 mRNA 的 Poly(A) 末端。
4、等温扩增实验中的组份。
活性定义
以 40 pmol 荧光标记的 25 bp RNA:DNA 杂交体为底物，在 50 μl反应体系中，50°C条件下，20 分钟内产生1nmol 核糖核苷酸所需的酶量定义为1个活性单位(U)。
质量控制
1、非特异性内切酶活性检测
将5 U的本酶与超螺旋质粒 DNA 在 37°C温育 4 h，通过 DNA 电泳检测质粒无变化。
2、DNase 残留检测
将5 U本酶与双链 DNA 底物在 37°℃温育 16 h，通过 DNA 电泳检测双链 DNA 底物无变化。
3、RNase 残留检测
将5 U的本酶与 500 ng RNA 在 37°C温育1h，使用琼脂糖凝胶电泳检测超过 90% 的 RNA 仍保持完整。
4、宿主 DNA 残留检测
本品中大肠杆菌宿主细胞 DNA 残留量低于 10 拷贝 /5 U。
注意事项
1、Thermostable RNase H反应缓冲液包含 MgCl2。目标 RNA:DNA杂交体和其它单链 RNA(总 RNA)在高温下使用 ThermostableRNase H时，建议控制反应时间和温度，以降低金属离子的单链RNA 的降解。
2、Thermostable RNase H可以通过添加蛋白酶K或过量的 EDTA来失活。
3、最佳反应温度 >65°℃，最高 95°℃。Thermostable RNase H在 65°C时的活性是 37°C时的 3 到 4倍。4.为了您的安全和健康，请穿实验服并带一次性手套操作。
| T751065 | Component | 200 U | 1000 U | Storage | | T751065A | Thermostable RNase H | 200 U | 5×200 U | -20°C. Avoid freeze/thaw cycle | | T751065B | 10× RNase H Reaction Buffer | 1 mL | 5×1 mL | -20°C. Avoid freeze/thaw cycle |
|
Thermostable RNase H (Thermostable RNase H) is an endoribonuclease that remains active at higher temperatures (above 65°C). Thermostable RNase H selectively recognizes and enzymatically cuts RNA strands in a heterozygous double strand of DNA, while remaining intact in the heterozygous double strand. ThermostableRNase H does not degrade single - or double-stranded RNA or DNA.
Product application
1. Preparation of high-precision RNA structure map and analysis of mRNA capping rate.
2. mRNA was removed from second strand synthesis of CDNA; Improve RT-qPCR efficiency.
3. Remove the Poly(A) end of mRNA in the presence of Oligo(dT).
4. Components in isothermal amplification experiment.
Activity definition
The amount of enzyme required to produce 1nmol ribonucleotides within 20 minutes in a 50 μl reaction system at 50°C using a 40 pmol fluorescently labeled 25 bp RNA:DNA hybrid as substrate was defined as 1 unit of activity (U).
Quality control
1. Detection of non-specific endonuclease activity
5 U of this enzyme was incubated with superhelix plasmid DNA at 37°C for 4 h, and no changes were detected by DNA electrophoresis.
2. DNase residue detection
The enzyme was incubated with double-stranded DNA substrate at 37° C for 16 h, and the double-stranded DNA substrate was detected by DNA electrophoresis.
3. RNase residue detection
After 5 U of this enzyme was incubated with 500 ng RNA at 37°C for 1h, more than 90% of the RNA remained intact by agarose gel electrophoresis.
4. Host DNA residue detection
The residual amount of Escherichia coli host cell DNA in this product is less than 10 copies /5 U.
Attention
1.Thermostable RNase H reaction buffer contains MgCl2. Target Rnas: When DNA hybrids and other single-stranded Rnas (total Rnas) are used with ThermostableRNase H at high temperatures, it is recommended to control the reaction time and temperature to reduce the degradation of single-stranded Rnas of metal ions.
2.Thermostable RNase H can be inactivated by adding protease K or excessive EDTA.
3. The best reaction temperature is >65° C, the highest is 95° C. Thermostable RNase H is three to four times more active at 65°C than at 37°C. 4. For your safety and health, please wear a lab coat and wear disposable gloves.
ZJ25F0319679
危险品化学品经营许可证（带存储）