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- 详细信息
- 文献和实验
- 技术资料
- 库存：
大量
- 英文名：
SgeI (3 U/µL)
- 保质期：
1年
- 供应商：
上海沪震实业有限公司
- 保存条件：
2-8℃保存
- 规格：
250units
SgeI (3 U/µL)
Enzyme : SgeI保存温度 : -20℃
货期 : 2-3天 Compatible
Buffer : Unique Buffer (10x Buffer SgeI) Optimal Reaction
Temperature : 37° C Sensitive to Heat
Inactivation: : Yes Methylation Sensitivity : Not CpG methylation-sensitive, Not dcm methylation-sensitive
5' Cm5 N N G N9 ↓ 3'
3' G N N C N13 ↑ 5' SgeI cleaves DNA targets containing 5-methylcytosine on one or both DNA strands
Thermo Scientific SgeI restriction enzyme recognizes m5CNNG(9/13)^ sites and cuts best at 37°C in its own unique buffer. See Reaction Conditions for Restriction Enzymes for a table of enzyme activity, conditions for double digestion, and heat inactivation for this and other restriction enzymes.
Thermo Scientific conventional restriction endonucleases are a large collection of high quality restriction enzymes, optimized to work in one of the buffers of the Five Buffer System. In addition, the universal Tango buffer is provided for convenience in double digestions. All of the enzymes exhibit 100% activity in the recommended buffer and reaction conditions. To ensure consistent performance, Thermo Scientific restriction enzyme reaction buffers contain premixed BSA, which enhances the stability of many enzymes and binds contaminants that may be present in DNA preparations.
Features
• Superior quality—stringent quality control and industry leading manufacturing process
• Convenient color-coded Five Buffer System
• Includes universal Tango buffer for double-digestions
• BSA premixed in reaction buffers
• Wide selection of restriction endonuclease specificities
Applications
• Molecular cloning
• Restriction site mapping
• Genotyping
• Southern blotting
• Restriction fragment length polymorphism (RFLP)
• SNP
Note: DNA methylated by Dcm or CpG methyltransferases will be a substrate for SgeI. Greater than 3-fold overdigestion with SgeI may result in incomplete cleavage. At least two copies of SgeI recognition sequence are required for an efficient cleavage. Amount of the enzyme required for complete digestion of methylated DNA depends on the number of SgeI recognition sites. DNA cleavage products generated by target site cleavage facilitate the nonspecific cleavage by SgeI. Therefore, optimization of the enzyme amount is recommended for DNA cleavage. pBR322 DNA isolated from E. coli dcm+ strain (#SD0041) can be used as a DNA cleavage efficiency control. SgeI cleaves all six dcm methylated targets on pBR322 DNA. For methylation sensitivity, refer to product specifications.
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文献和实验Single Primer ("Semi-Random") P
to about 1 ng/µL 3 µL "T/TS" @ 0.4 u Taq/µL -------- 30 µL Cycle as follows: 30"x94°C 20 cycles 0"x94°C 0"x55°C 1'x72° S=9 30 cycles 0"x94°C 0"x40°C 1'x72° S=6 30 cycles 0"x94°C 0"x55°C 1'x72° S=9 Clean up: Add 1 µL ExoI
x PCR reaction buffer, 1.6 µl 4dNTP mix (250 pmol/µl), 2 µl T12 MN primer, 2 µl arbitrary primer (decamer), 0.2 µl Taq DNA polymerase (5 U/µl) use the PCR conditions as under section III. run PCR product on a 1.5 % agarose gel
Single tube confirmation PCR protocol
. For example, add 5 µl of the Zymo solution from isolate #1 to PCR tubes 1-5, 5 µl of the Zymo solution from isolate #2 to PCR tubes 6-10 and so on. - Make a PCR master mix by combining: 638 µl water, 110 µl 10 x Taq Buffer, 11 µl NTP's, and 11 µl Taq
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