CIAP(小牛肠碱性磷酸酶),1 U/μL
CIAP(小牛肠碱性磷酸酶),1 U/μL
Invitrogen™

CIAP(小牛肠碱性磷酸酶),1 U/μL

Calf Intestinal Alkaline Phosphatase (CIAP) is a phosphomonoesterase that removes 3´ and 5´ phosphates from DNA and RNA.Concentration: 1 unit/μLApplications:了解更多信息
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货号数量
180090271000 U
货号 18009027
价格(CNY)
660.00
Each
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数量:
1000 U
请求批量或定制报价
价格(CNY)
660.00
Each
添加至购物车
Calf Intestinal Alkaline Phosphatase (CIAP) is a phosphomonoesterase that removes 3´ and 5´ phosphates from DNA and RNA.

Concentration: 1 unit/μL

Applications:
Dephosphorylation of 5´-phosphorylated termini of vector DNA to prevent self-ligation (1). Dephosphorylation of 5´ termini of nucleic acids prior to forward reaction with kinase.

Source: Purified from calf intestinal mucosa.

Performance and Quality Testing:
Endodeoxyribonuclease, 3´ exodeoxyribonuclease, and ribonuclease assays; dephosphorylation efficiency measured in a transformation assay.

Unit Definition:
One unit hydrolyzes 1 μmol of 4-nitrophenyl phosphate in 1 min. at 37°C.

Unit Reaction Conditions:
1 M diethanolamine buffer, 10 mM 4-nitrophenyl phosphate, 0.25 mM MgCl2 (pH 9.8) in 900 μl for 10 min. at 37°C.
仅供科研使用。不可用于诊断程序。
规格
兼容缓冲液二乙醇胺缓冲液
产品类型碱性磷酸酶
数量1000 U
运输条件经批准可置于湿冰或干冰上运输
最大浓度1 U/μL
CIP
Unit SizeEach
内容与储存
小牛肠碱性磷酸酶(1 单位/µL)配备一瓶 10X 去磷酸化缓冲液 [500 mM Tris-HCl(pH 值 8.5)、1 mM EDTA] 和一瓶稀释缓冲液 [50% 甘油、25 mM Tris-HCl(pH 值 7.6)、1 mM MgCl2 和 0.1 mM ZnCl2]。小牛肠碱性磷酸酶储存在 -20°C 下。

常见问题解答 (FAQ)

进行限制性内切酶克隆除了连接酶以外还需要什么酶?

对载体进行去磷酸化以降低背景时可以使用:
热失活的碱性磷酸酶,或
牛小肠碱性磷酸酶(提供1 unit/μL或20 units/μL)规格)

如果需要获得磷酸化的DNA平末端(“补平”末端),您可以使用:
可以用DNA End Repair Mix或T4 DNA 聚合酶或E. coli DNA聚合酶Klenow片段(大片段)来产生平末端,因为它们有5’ →3’DNA聚合酶活性(补齐5’突出端)和3’→5’外切酶活性(切除3’突出端)。

去除5’磷酸基团,应使用什么酶?添加磷酸基团,应使用什么酶?

CIP/CIAP 或BAP可以对DNA/RNA的5’端去磷酸化;T4多核苷酸激酶可以添加磷酸基团。

What other enzymes besides ligase may I need to perform restriction cloning?

Dephosphorylating the vector to decrease background can be achieved with:
Heat Inactivated Alkaline Phosphatase, or Calf Intestinal Alkalline Phosphatase (available as 1 unit/µL or 20 units/µL)

If you need to create blunt phosphorylated DNA ends (“polishing” the ends), you can use:
DNA End Repair Mix or T4 DNA polymerase or Klenow Fragment (large fragment) of E. coli DNA polymerase to generate blunt ends due to their 5' to 3' DNA polymerase activity (filling-in of 5' overhangs) and 3' to 5' exonuclease activity (chewing back of 3' overhangs).

What enzymes can be used to remove 5' phosphate groups, and what enzyme can be used to add the phosphate groups back on?

CIP/CIAP or BAP can dephosphorylate the 5' ends of DNA/RNA. T4 Polynucleotide Kinase can add back phosphate groups.

Why are alkaline phosphatases used in cloning protocols? What is a typical protocol for dephosphorylation of nucleic acids?

Alkaline phosphatases are used to dephosphorylate the 5' ends of DNA. In cloning, it is used to prevent self-ligation of vector DNA. Standard ligation of DNA with ligase requires a 5' phosphate to be present on at least one of the ends being joined. When a DNA insert containing phosphates on both 5' termini is added to a dephosphorylated vector, the insert will be efficiently ligated into the vector, but the vector will not be able to self-ligate. Thus, dephosphorylation of vector lowers the number of background colonies containing vector without insert.