pLUG-Prime® TA-cloning Vector Kit II
型号:
价格:联系人：王,请致电：010-67529703,手机(微信同)：18210996806,QQ:997554969
,品牌:intronbio
pLUG-Prime® TA-cloning Vector Kit II
Description
Fast ligation and high transformation efficiency pLUG-Prime® TA-Cloning Vector Kit II
- • Possible to insert DNA ligation of various sizes in a short time of 5-15 minutes
- • High true white colony ratio Blue / white colony selection function
- • Selective culture with ampicillin resistant gene
- • Use M13 primer site for convenient sequencing
- • Application of highly restrictive enzymes
- • EcoRI restriction enzyme site is on both sides of MCS (Multi cloning site), enabling enzyme cutting at one time
pLUG-Prime® TA-cloning Vector Kit II features fast ligation and high transformation efficiency. Especially, it is designed to ligation within 5-15 minutes. It shows high true white colony ratio in various size insert DNA. It is also possible to cultivate selectively using ampicillin resistant gene. Blue / White colony sorting function makes it easier to select target transformants. The pLUG-Prime® TA-cloning Vector Kit II TA-cloning site is more common and is designed to have a well-known restriction enzyme site on which, in particular, its predecessor pLUG-Prime® TA-cloning EcoRI restriction enzyme site relative to both Vector Kit This method makes it easy to carry out restriction analysis of recombinant plasmids and re-cloning into other vectors.
Applications
- 01TA-cloning
- 02 Accepts terminal 3’-dA nucleotides overhang PCR products
- 03Transform ligation product (purified/unpurified) into competent cells
- 04LacZ complementation for blue/white screening
- 05 Ampicillin marker for antibiotic selection
Kit Contents
| Content | Concentration | Volumes |
|---|---|---|
| TA-cloning Vector II (20 reactions) | 25 ng/μl | 40 μl × 1 vial |
| Control insert DNA | 10 ng/μl | 10 μl × 1 vial |
| T4 DNA ligase | 2 U/μl | 20 μl × 1 vial |
| Ligation Buffer A | - | 50 μl × 1 vial |
| Ligation Buffer B | - | 50 μl × 1 vial |
| Forward Primer (M13-F) | 10 μM | 50 μl × 1 vial |
| Reverse Primer (M13-R) | 10 μM | 50 μl × 1 vial |
| Manual | 1 ea |
Technical Data
Map & Multiple cloning site of the pLUG-Prime® TA-cloning vector Kit
Figure 1 : Map and sequence reference points of the pLUG-Prime® TA-Cloning Vector II
* Before the insert is incorporated into the pLUG-Prime® TA-Cloning Vector II, there is only one HindⅢ site and no BglⅡ site. After the incorporation, the T and A nucleotide on the insert will complement with the sequence on the vector and generate these two new sites.
This merit of pLUG-Prime® TA-Cloning Vector II makes cloning more economical and convenient.
Figure 2 : Multiple cloning site sequence of the pLUG-Prime® TA-Cloning Vector II
Figure 3. The gel analysis of the PCR products ligated by pLUG-Prime® TA-Cloning Vector Kit II
- QIn the manual, PCR products are supposed to be used 5-10 times the amount of vector. How many ng per 1ul vector?
- AThe concentration of the vector provided in the kit is 25 ng / ul. The total concentration should be 50 ng, since 2 μl should be used per experiment. For more details on the amount of PCR products used, please refer to the "Optimizing insert to vector ratio" section of the manual.
- QAfter the transformation, the number of colony is high but the ratio of blue colony is higher.
- ABlue colony may vary, but be sure to check that all components are properly placed when ligation first occurs. Another case is when the bacteria strain is LacZ- (LacZ mutant strain) at the time of transformation. It is recommended that you check the strain phenotype before the experiment.
- QThe size of PCR product to be used as Insert DNA is 4.5Kb. Is it OK to carry out the ligation time for about 15 minutes for the PCR product of this size?
- AThis product is available for fast ligation. In case of insert DNA of general size, ligation time is about 5-15 minutes. However, large insert DNA, such as 4.5Kb product, is recommended to last a little longer. Therefore, if you want to obtain high efficiency test results with large insert DNA of 4.5Kb, we recommend ligation reaction at room temperature for more than 8 hours.
- QpLUG-Prime® TA-Cloning Vector Kit II for cloning and sequencing. What sequencing primer should be used?
- AAs you can see from the pLUG-Prime ® TA vector II map, you can use the M13 primer.
- QIs Blue/White colony selection possible?
- AYes. It is possible. As you can see from the map of Vector, our products are designed with LacZ based.
- QWhen previous samples were grown in the medium, ampicillin and kanamycin were used as antibiotics. The pLUG-Prime® TA-Cloning Vector Kit II contains only the ampicillin resistant gene. I already finished smelling it. Will Colony be asleep?
- AWe recommend you do the experiment again. In our case, the colony will not be produced because it is not kanamycin resistant.
- QWe want to obtain insert DNA using only one restriction enzyme. Is there any other way to use the M13 primer?
- AYes, it is possible. As you can see from the map of this vector, Eco RI restriction enzyme exists at both ends of MCS (multi cloning site). Therefore, vector and insert DNA can be separated using Eco RI restriction enzyme only.
| Product | Cat.No | Capacity | inquire | |||||||
|---|---|---|---|---|---|---|---|---|---|---|
| MacCell™ DH5α (10^7) |
| |||||||||
| MacCell™ DH5α (10^8) |
| |||||||||
| Maxime™ PCR PreMix (i-Taq) |
| |||||||||
| Maxime™ PCR PreMix (i-StarTaq) |
| |||||||||
| Maxime™ PCR PreMix (i-MAXⅡ) |
| |||||||||
| DNA-midi™ GT Plasmid DNA Purification Kit |
| |||||||||