图2 The helper cell should meet two basic requirements:(1)it should provide functions missing from the the vector virus,and(2)it should not be capable of producing viable virus particles. The crucial element in the development of a successful helper cell is the design of the helper virus. Researchers use recombinant DNA techniques to disable the helper virus in the test tube-one of the most common measures is removal of the Psi fragment. The Psi-deficient helper virus produces all of the normal viral proteins, but cannot package its own RNA because it lacks the appropriate packaging signal. Helper virus DNA is inserted into the genome of the helper cell using chemical techniques.
STEP 3
PRODUCING THE VECTOR
图3 Scientists use chemical measures or an infection technique to insert recombinant vector DNA (including a human gene) into helper cells. Because the vector provirus contains the Psi sequence, the vector RNA genome is automatically encapsulated by viral proteins produced by helper virus DNA in the helper cell. The resulting viral particles are released by budding from the helper cell membrane. The vector virus is capable of only one infection because it lacks the information needed to make viral proteins.
STEP 4
INFECTING THE TARGET ELL
图4 Researchers infect target cells (such as human bone marrow cells) with the vector virus in two different ways: they mix them with helper cells producing the virus, or they bathe them in fluid harvested from the helper cell culture. When the vector provirus is integrated into the target cell DNA ,enzymes from the target cell treat it as an integral part of the genome. Cellular enzymes do the work necessary to make proteins from the foreign genes located between the two viral L TRs.
