BioPioneer’s unique high-throughput format competent cells are arrayed in breakable 24-well, 48-well and 96-well PCR plate.The plates can be separated without scissors into any combination of 8 well strips. DH5alpha is one of the most commonly used E. coli strain for plasmid transformation. Ultra-high efficiency DH5alpha Competent Cells are prepared with unique salt compositions and procedures that result in significantly higher transformation efficiency than those by traditional lab protocols using CaCl2 buffers. Biopioneer’s competent cells are quality controlled by direct comparison to other leading brands of similar products in transformation of ligated plasmids.
Ultra-high efficiency DH5α Competent Cells are suitable for propagation of plasmids of all sizes and preparations.They are recommended for difficult constructs that often yield low colony counts.
Features:
- Widely used strain
- Suitable for 3 min quick protocol
- Efficiency: Constantly above 1 x 109 cfu/µg of supercoiled pUC19 plasmid DNA; higher colony counts than DH5a Competent Cells from other leading suppliers, especially when transforming ligation reactions.
- Conveniently packaged in 60ul aliquot, no repeated freezing in dry ice and alcohol bath.
- Blue/white color screening of recombinants
Genotype:
F’/endA1 hsdR17(rK-mK+) supE44 thi-1 recA1 gyrA (Nalr) relA1 D(laclZYA-argF)U169 deoR
(F80dlacD(lacZ)M15)
Storage:
It has been observed that long term storage of competent cells at -80oC can actually cause the cells to adapt by increasing the recombination rate. Therefore, BioPioneer Inc. prepares its competent cells frequently at small scales. It is recommended that competent cells be used within 3 months after arrival.
Transformation Protocol
1. Thaw competent cells on ice. It is essential that cells be completely thawed before pipetting. Pipet slowly.
2. Add DNA of your choice (pure DNA or ligation mix) to the cells and incubate on ice for 30 minutes (may use 5 min for subcloning). Heat-shock at 42°C for 30 seconds and put the tube on ice for 1.5-2 minutes.
3. Add 200ul SOC or LB broth and shake the cells at 37°C at 225 rpm for 30-60 minutes.
4. Plate out desired amount on a prewarmed (37°C) plate with the appropriate antibiotic and incubate in a 37oC incubator.
Quick Protocol:
1. For each transformation, thaw one tube of cells on ice.
2. Mix DNA and cells, incubate on ice for 1-2 min
3. Heat shock in 42°C water bath for 45 sec, or in 37°C water bath for 1.5 min
4. After heat-shock, incubate the reaction on ice for 1 min and spread on plate directly.
The typical 1 hour growing in plain LB may generate 2-3 fold more colonies but is not necessary in most cases.
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看有关产前诊断的一个资料，提到：“凡是酶活性能在成纤维细胞中表达均可用绒毛或羊水细胞的酶活性测定进行产前诊断，如溶酶体贮积症。”
不太理解，为什么成纤维细胞中表达的酶就能在绒毛或羊水细胞中测出来呢？绒毛的细胞跟成纤维细胞没什么直接关系吧？或者是因为成纤维细胞脱落到羊水中，破裂，然后就可以检测了？
求各位解答，谢谢。
如癌症的CA抗原系列、用于测胆汁酸、尿酸之类的酶。
转换酶中的一类。催化氨基酸和a-氧代酸（a-酮酸
)或醛酸之间的氨基转换反应，生成与原来的a-氧代酸或醛酸相应的a-氨基酸或ω-
氨基酸，原来氨基酸转变成相应的氧代酸。转氨酶催化的反应都是可逆的。转氨酶可按底物的不同分成3大类。L-a-氨基酸（酮酸转氨酶） 、ω-
氨基酸（酮酸转氨酶）和D-氨基酸转氨酶。转氨酶的辅基是磷酸吡哆醛或磷酸吡哆胺，两者在转氨基反应中可互相变换。
转氨酶参与氨基酸的分解和合成。氨基酸转氨后生成的酮酸或醛酸可经氧化分解而供能，也可转变成糖类或脂肪酸。相反，酮酸或醛酸也可经转氨酶的作用而生成非必需氨基酸。某些氨基酸之间的互变也有转氨酶参与。在高等动物各组织中，活力最高的转氨酶是谷氨酸
： 草酰乙酸转氨酶（ GOT ）和谷氨酸：丙酮酸转氨酶（GPT）。
转氨酶是人体代谢过程中必不可少的“催化剂”，主要存在于肝细胞内。当肝细胞发生炎症、坏死、中毒等，造成肝细胞受损时，转氨酶便会释放到血液里，使血清转氨酶升高。
通常，体检中主要检查的转氨酶有丙氨酸转氨酶（ALT，俗称谷丙转氨酶）和天门冬氨酸转氨酶（AST，俗称谷草转氨酶），其中尤以前者（ALT）最为常用。1%的肝脏细胞损害，可以使血中ALT的浓度增加1倍。因此，ALT水平可以比较敏感地监测到肝脏是否受到损害。
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