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More information about the genes added to HoneySweet—

Were other new genes added?
In addition to the coat protein from the PPV, two other genes—beta glucuronidase (GUS) and neomycin phosphotransferase (NPTII)—were added in the same piece of DNA that carried the coat protein to provide markers that allow researchers to easily track the presence or absence of the new DNA. GUS is a gene that produces an enzyme that will react with an indicator chemical (X-Glu) by turning it blue, demonstrating that the tissue has the new genes in it. The NPTII gene produces an enzyme that makes the tissue resistant to the antibiotic kanamycin. If tissue grows in the presence of kanamycin, we know the new genes are actually in the cell—including the PPV coat protein gene since it is located next to the NPTII gene in the carrier DNA.

Does the tree or fruit contain antibiotics? Are these harmful to people or animals eating the fruit or leaves?
The fruit and leaves do not contain any antibiotics, only the antibiotic resistance gene. In addition, it is possible that the leaves and fruit may contain a small amount of the enzyme produced by the NPTII gene that produces kanamycin resistance. This gene and the enzyme it produces have been approved by the FDA in 1994 as safe to be present in food.

Does the resistant tree make the virus coat protein?
No.

If it does not make protein, what makes the tree resistant?
The resistance is due to RNA silencing, which is a natural process in plants that gives them some adaptive protection against viruses. In the silencing process, the introduced PPV coat protein gene induces the plant to make an enzyme that breaks down the coat protein, which prevents virus infection. This is a natural mechanism that plants use not only for virus resistance, but also to regulate many normal cell processes.

Is there other transgenic fruit that has been made by adding virus coat protein DNA?
Yes. Papaya has been genetically engineered to resist ringspot virus by adding a virus coat protein. These papayas are being grown in Hawaii and sold there and in the U.S. mainland and Canada. Squash has also been genetically engineered with a virus coat protein.

Could a PPV-resistant tree be developed through conventional breeding?
A few highly resistant stone fruit or ornamental stone fruit varieties have been developed through conventional breeding. Hybrid plum cultivars have been identified that respond to PPV by a hypersensitive response. This means that once virus infection occurs, the plant tissue surrounding the infection site quickly dies. PPV, like all other viruses, can survive and multiply only in living host cells. Therefore, the hypersensitive response prevents spread of the virus to other parts of the tree and eliminates the virus from the host.

Unfortunately, the hypersensitive response in these plum hybrids is regulated by several different genes in the tree, making it difficult and time-consuming to incorporate the hypersensitive trait into other cultivars by standard plant breeding methods.

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