| 摘要 |
A method of introducing a target gene sequence into a primed algal cell, comprising a step of providing a primed algal cell comprising providing an algal cell; introducing into the algal cell an integration cassette comprising at least two site-specific recombination sites and at least one selectable marker gene wherein the at least two site-specific recombination sites are positioned to flank the at least one selectable marker gene; and selecting cells which have incorporated the integration cassette by cultivating the cells in a selective media and selecting growing cells, wherein an ability of the cells to be cultured on the selective media is dependent on a presence of the at least one selectable marker in a genome of the algal cell. The site-specific recombination sites may be compatible. The method includes a further step of effecting targeted site-specific recombinase mediated deletion of the target cassette can be carried out. The site-specific recombination sites maybe heterospecific to permit introduction of target gene(s). The method further comprises a step of providing a target cassette comprising at least one target gene sequence flanked by a type I site-specific recombination site and a type II site-specific recombination site, these sites being capable of recombining with those in the primed algal cell. Moreover, the method includes a further step of effecting targeted site-specific recombinase mediated insertion of the target cassette into the algal genome by effecting recombination between corresponding type I and type II site-specific recombination sites flanking the target gene sequence and located in the algal genome, such that the target gene sequence is introduced into the algal genome replacing the at least one selectable marker. The invention also concerns integration cassettes for use in the method above, as well as an algal cell for use with and modified algal cell produced by the above method. |
