After four years of attempting to trace hosta ancestry through DNA fingerprinting, Dave Koetje has come to a conclusion: “Hostas are weird,” he says.

Since 2002 Koetje, a Calvin College biology professor and director of the college’s biotechnology program, along with various student researchers, have been trying to track hosta parentage genetically, research that is funded by the and Calvin’s science division and biology department.

The project is a collaboration with , a Zeeland-based perennial plant wholesaler, which grows its many varieties of hostas in tissue culture.

Koetje describes the process: “You take a little snip, and you plant it in the test tube or jar that has the growth medium in it. It’s not that different from how people usually propagate hostas. You’ve got these hosta plants growing in your yard, and your neighbor comes over and says, ‘Wow! That’s a pretty one.’ And you say, ‘I’ve got to split it anyway.’ And you split it with a shovel.”

Walter’s Gardens expedites this process, he says, by propagating up to 50 hostas in one Mason jar from little snippets of plant material, using growth hormone.

“They’ll grow them, transplant them into a pot, transfer them into a greenhouse to raise them, and from there they sell them to a place like Flowerland," he says.

Hostas, whether grown in nature or in tissue culture regularly produce mutations or “sports.”

Koetje notes that the mutant might have a new variegation pattern, or it might have some speckles on it, something that makes it stand out. The plant’s mutating tendency accounts for some 4500 varieties of hostas that have descended from a mere three or four hosta species.

“There are people who pay big bucks for the latest and greatest varieties,” Koetje says.

Plant purveyors typically patent their new varieties, but it is difficult to distinguish a sport grown in one nursery from a sport grown in another.

Clarence Falstad, the tissue culture lab manager at Walter’s Gardens (and a “hosta guru” according to Koetje), asked for help in determining hosta parentage genetically — to establish a hosta family tree — and the American Hosta Society contributed $5,500 to fund the project.

Koetje set out to determine how hostas are related to one another using Amplified Fragment Length Polymorphism (AFLP) or DNA fingerprinting.

“You extract the DNA, cut it, make copies of it, separate the copies by size and from that construct a DNA fingerprint,” Koetje explains.

At first, he and student researchers Sarah Weeda and Jody Mohle struggled because of low-quality DNA samples from hostas collected in the fall. Once they started using fresher hosta DNA, they encountered new complications: It seemed impossible to get a reliable DNA fingerprint from a hosta plant.

The team had to decide if this was a procedural glitch or did these plants have cells in them with slightly different amounts of DNA.

"After months of procedural optimization," says Koteje, "we gradually came to the conclusion that the DNA in hosta can vary from leaf to leaf in a single plant. People have known for years that some plants have different ploidy levels — different amounts of DNA in them. And yet the plant is just fine. Now, our bodies would freak out if that happened.”

He also found a parent hosta and its sport could have very different genetic makeup, a conclusion confirmed by the research of his colleague, John Beebe.

Beebe, who has taught biology at Calvin for so long that Koetje was once his student researcher, and Vonny Salim, his current student researcher, are using a flow cytometer (chromosomes are stained with a fluorescent compound, then the cells are passed single-file in front of a detector) to measure the amount of DNA in the cell nuclei of two hosta varieties: Regal Splendor and its parent plant, Krossa Regal.

“Regal Splendor has twice the amount of DNA as Krossa Regal,” says Beebe. “There’s a lot more diversity than you’d expect.”

Koetje agrees.

“We think that what happens is that there is a wholesale chromosome rearrangement when a sport arises in culture,” he says. “It’s like a bomb goes off in a nucleus. It reminds us of what they’re discovering with cancer, but in this case, you don’t end up with runaway cell growth. You end up with slight changes in leaf pattern.”

By growing hostas in tissue culture Walter’s Garden’s inadvertently accelerates the mutating process.

“It’s happening more frequently in tissue culture in part because you’re doping them up on some hormones, and that makes the cells divide quickly, and when they divide quickly, they make genetic mistakes, Koetje says. “It’s not entirely natural, but it’s not entirely unnatural either.”

He approached his hosta research, Koetje says, thinking that the mutations that occur to produce a sport would be so small that he wouldn’t be able to measure them only to find out the opposite was true.

“There’s such a big genetic change," he says, "I can’t tell if parents and children are related to each other.”

Koetje is still examining the procedure behind his research for possible glitches, even as he gradually concludes that the hosta family tree may be impossible to reconstruct.

“Practically, it doesn’t help the hosta industry at all,” he says. “Genetically it’s a very interesting story. The biology of this turns out to be interesting and the practical stuff turns out to be a dead-end street. The plant kingdom has some surprises."

Beebe, a fan of hostas sees the benefit for a business like Walter’s Garden’s.

“They have a fantastic catalogue,” he says.