그림 1. In-Fusion Snap Assembly와 In-Fusion HD (기존 제품) 간의 효율 비교
In-Fusion Snap Assembly는 기존 제품에 비해 정확도는 유지하면서, colony 형성 능력이 개선되었다.
Five different inserts (ranging in size from 405 bp to 1,005 bp) were simultaneously cloned into a 2.7-kb vector previously linearized by inverse PCR using In-Fusion HD and In-Fusion Snap Assembly. Each cloning reaction was performed in triplicate. Amplification primers and all reagents, except for the cloning master mixes, were the same between the two systems. After transformation and plating, 10 colonies from each replicate were analyzed by Sanger sequencing to determine the cloning accuracy. In-Fusion Snap Assembly yielded 4X more colonies than In-Fusion HD while maintaining high cloning accuracy.
그림 2. In-Fusion Snap Assembly와 NEBuilder HiFi (1)
PCR로 선형화한 vector에 single insert를 cloning한 결과, insert 크기에 관계없이 NEBuilder HiFi보다 In-Fusion Snap Assembly에서 훨씬 많은 colony 수를 보였다. 정확도는 95% 이상이었다.
A single 3.8-kb insert (Panel A) or a 34.2-kb adenovirus insert (Panel B) was cloned into a 2.7-kb vector which was linearized via inverse PCR. These cloning reactions were performed in triplicate with both In-Fusion Snap Assembly and NEBuilder HiFi. Primers were designed according to the manufacturers' specifications. After transformation and plating, 20 colonies from each replicate were analyzed by Sanger sequencing (for the 3.8-kb insert) or colony PCR (for the adenovirus insert) to determine the cloning accuracy. In-Fusion Snap Assembly yielded 2X more colonies than NEBuilder HiFi.
그림 3. In-Fusion Snap Assembly와 NEBuilder HiFi (2)
제한효소로 선형화한 vector에 single insert를 cloning한 결과, 말단의 형태 (overhangs, blunt)에 관계없이 NEBuilder HiFi보다 In-Fusion Snap Assembly에서 훨씬 많은 colony 수를 보였다.
A single 3.8-kb insert was cloned into a standard cloning vector linearized by restriction digests to create 5' overhangs (Panel A), blunt ends (Panel B), or 3' overhangs (Panel C). These cloning reactions were performed in triplicate with both In-Fusion Snap Assembly and NEBuilder HiFi. Primers were designed according to the manufacturers' specifications. After transformation and plating, 20 colonies from each replicate were analyzed by Sanger sequencing to determine the cloning accuracy. In-Fusion Snap Assembly yielded between 5X and 16X more colonies than NEBuilder HiFi, depending on the type of linearized vector ends.
