Vũ Hải Chi
Senior Member
Không biết có bác nào dùng kit này chưa nhỉ, chẳng biết hiệu quả thực hư ra sao
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The GeneJET™ PCR Cloning Kit is an advanced positive selection system for the highest efficiency cloning of PCR products generated with any DNA polymerase. The kit offers cloning efficiencies of up to 100% positive clones, eliminating the need for tedious colony screening. The kit is specifically designed to ligate PCR products into a positive selection vector in just 5 minutes. Purification of PCR products is not required.
Blunt-end PCR products generated with a proofreading DNA polymerase (e.g., Pfu DNA polymerase) are directly ligated into the cloning vector. PCR products generated either with non-proofreading DNA polymerases (e.g., Taq DNA polymerase) or with mixtures of DNA polymerases are blunted in 5 minutes prior to ligation.
All common laboratory E.coli strains can be directly transformed with the ligation product*. The recommended efficiencies of competent cells are 107cfu/µg and higher. This level of transformation efficiency is easily achieved with the TransformAid™ Bacterial Transformation Kit.
The GeneJET™ PCR Cloning Kit includes a novel, ready-to-use positive selection cloning vector pJET1/blunt (see Fig.1). pJET1/blunt is a positive selection vector, therefore only recombinant plasmids containing the insert DNA are propagated after transformation into E.coli. Recircularized pJET1/blunt vector molecules lacking the insert express a lethal restriction enzyme after transformation, killing the host E.coli cell. The positive selection drastically accelerates the process of colony screening and eliminates additional costs of expensive reagents IPTG and X-Gal required for cloning into vectors with blue/white selection.
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_h2tp://www.fermentas.com/catalog/kits/genejetpcrclon.htm
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The GeneJET™ PCR Cloning Kit is an advanced positive selection system for the highest efficiency cloning of PCR products generated with any DNA polymerase. The kit offers cloning efficiencies of up to 100% positive clones, eliminating the need for tedious colony screening. The kit is specifically designed to ligate PCR products into a positive selection vector in just 5 minutes. Purification of PCR products is not required.
Blunt-end PCR products generated with a proofreading DNA polymerase (e.g., Pfu DNA polymerase) are directly ligated into the cloning vector. PCR products generated either with non-proofreading DNA polymerases (e.g., Taq DNA polymerase) or with mixtures of DNA polymerases are blunted in 5 minutes prior to ligation.
All common laboratory E.coli strains can be directly transformed with the ligation product*. The recommended efficiencies of competent cells are 107cfu/µg and higher. This level of transformation efficiency is easily achieved with the TransformAid™ Bacterial Transformation Kit.
The GeneJET™ PCR Cloning Kit includes a novel, ready-to-use positive selection cloning vector pJET1/blunt (see Fig.1). pJET1/blunt is a positive selection vector, therefore only recombinant plasmids containing the insert DNA are propagated after transformation into E.coli. Recircularized pJET1/blunt vector molecules lacking the insert express a lethal restriction enzyme after transformation, killing the host E.coli cell. The positive selection drastically accelerates the process of colony screening and eliminates additional costs of expensive reagents IPTG and X-Gal required for cloning into vectors with blue/white selection.
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_h2tp://www.fermentas.com/catalog/kits/genejetpcrclon.htm
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nhưng trường hợp có rất ít khuẩn lạc trắng thì không phải là hiếm. Sử dụng kit xanh - trắng có thể giúp mình hoàn thiện kỹ thuật hơn. Chẳng hạn như lúc ligate lấy enzyme kô chuẩn (cảm giác), hoặc những sai sót nhầm lẫm nho nhỏ trong các thao tác, có thể tỉ lệ xanh-trắng sẽ giúp ta hình dung tốt hơn. Cái dở có lẽ là ở khâu trải X-gal vì ... ngại
