In vitro molecular directed evolution, based on the combination of random mutagenesis, recombination and high throughput screening, is a new strategy of improving protein properties. With this technology, the properties of the target proteins, such as specificity, catalytic activity and affinity can be tuned without the knowledge of 3-D structure information and function mechanisms. Recently, the methodology originating from error prone PCR and DNA shuffling has been greatly developed by many new techniques. Here, based on the literature review and the experience of authors, the recent developments have been reviewed.