Ollie, a Przewalski’s horse clone at five days old. (Elizabeth Arellano Photography)

Time Stands Still in San Diego’s Frozen Zoo

San Diego’s Frozen Zoo, where the living cells of critically endangered (and extinct) animals are safeguarded, might be one of the last lines of defense in the face of mass extinction

BEYOND SAN DIEGO’S suburbs, where ruffled lemon trees brimming with winter fruit give way to chaparral and rambunctious paddles of prickly pear, there exists a most curious zoo.

A zoo that freezes time.

Tanks pressurized at minus 320 degrees Fahrenheit line its walls. Described by its director as “neutron stars of biodiversity,” they’re filled with more than 11,000 deep-frozen living cell lines, eggs, sperm, and embryos—each frosty specimen cocooned in miniature, hand-labeled vials. By donning insulated gloves, you can plunge giant tweezers through sizzling liquid nitrogen to pull a critically endangered vaquita, or sea star, or blue-eyed black lemur straight out of the past.

San Diego’s Frozen Zoo was created by fertility scientist Dr. Kurt Benirschke in 1975, just two years after passage of the Endangered Species Act. Nearly fifty years later, what was once a laboratory curiosity has become the world’s largest living biobank of endangered plant and animal DNA. I’ve come to the Beckman Center for Conservation Research right before the new year to meet the Frozen Zoo’s curator, Marlys Houck, a cytogenetics specialist who runs the predominantly female team—an anomaly in a traditionally male field—that collects, grows, and preserves these cell lines. I have a deep fondness for specialists like these. After seven years of unexplained infertility and a round of in vitro, I’m intimately familiar with a few of the assisted reproduction technologies the Beckman Center supports for conservation. I know something about this particular kind of wild hope. As a new mother in the sudden quiet of the polished hallways, my thoughts curl around my daughter’s dimpled arms—she just started giving hugs. 

 

The Frozen Zoo preserves living cell lines, gametes, and embryos from more than 1,281 species and counting. (San Diego Zoo Wildlife Alliance) 
 

Marlys meets me in front of a window where we can view the Frozen Zoo. “It’s kind of a little nursery,” she says, smiling as together we peer through the glass. The cryotanks may as well be bassinets lined up in a mid-century hospital. This is not the cold, clinical setting I was expecting. It almost feels like a maternity ward.

Living cells arrive here from adult animals all over the world, usually taken as skin biopsies soon after death or during routine veterinary exams. “The trick is to stop the clock,” Marlys says. “The race, for us, is to not have the cells die before they get to the lab.” Cells can decompose. They can be contaminated by bacteria or fungi, perish from dehydration. Or, if the animal dies in the snow, frostbite can even puncture the delicate cell membranes. 

Back in the lab, these living skin tissues are processed and incubated to encourage growth of fibroblasts, a type of hardy, connective cell. A cryoprotectant is then added before they’re suspended in liquid nitrogen. The fresh DNA these fibroblasts hold can then be thawed decades later for genome sequencing—or cloning.

 

Marlys Houck is the Curator of San Diego Zoo Wildlife Alliance’s Frozen Zoo. (San Diego Zoo Wildlife Alliance)
 

This sounds like science fiction, I know, but just beyond the Beckman Center, a swarthy, Przewalski’s foal named Ollie gallops around a private enclosure. Ollie, like his identical twin, Kurt (named for the Frozen Zoo’s founder and born just three years prior), is a clone created from a stallion’s DNA banked more than forty years ago. 

Przewalski’s horses once ranged across Europe and Asia. Formerly extinct in the wild, herds today are descended from just twelve individuals: creating a severe genetic bottleneck. The plan is for Kurt and Ollie to be socialized by other captive Przewalski’s horses, then released into a wild herd as studs, infusing much-needed diversity and resilience back in to the genetic pool.

The word cloning can elicit strong reactions, but it does occur in nature—like budding in yeast, or fragmentation in worms, or parthenogenesis in tiger sharks. Even natural human clones can form when a fertilized egg splits in two, creating identical twins. Artificial clones like Ollie, however, are forged through a process called somatic cell nuclear transfer, where DNA from a donor egg (in this case, a domestic horse) is replaced with a full set of Przewalski’s horse chromosomes. The egg is then implanted into the womb of a surrogate. Ollie’s mother, a palomino quarter horse, delivered him at a genetics facility in Texas in early February 2023. 

 

Ollie, a 5-day-old Przewalski’s horse clone with his quarter horse mother. (Elizabeth Arellano Photography)
 

“He was up and nursing within a few minutes and only slows down for regular naps,” reported an excited collaborator at the birth announcement. 

Critics of cryoconservation cite inefficiencies and expense, and point out that it fails to address root causes of declines: degradation of habitat, poaching, and invasive species. Others, including the International Union of Conservation of Nature (IUCN), who recently partnered with San Diego’s Frozen Zoo to form a new Center for Species Survival, believe it represents just one of many useful conservation tools. 

“When I started here in the 1980s, I helped to bank northern white rhinos,” recalls Marlys. “At the time, they weren’t in any more danger than any other rhino—then they started to plummet.” With only two females still alive, and only one of breeding age, the twelve skin cell lines she helped bank contain invaluable DNA now integral to any kind of sustained recovery. 

“We weren’t able to do cloning when we started the Frozen Zoo,” she says. “What else will we be able to do in the future?” 

I know something about this particular kind of wild hope.

I catch myself—as so many new parents do—swaying back and forth as I listen. Marlys tells me the zoo safeguards living genetic material from three extinct species. That kind of responsibility, she admits, can make her lose sleep. 

“The po‘ouli, the Hawaiian songbird, that was a scary one.” When I ask why, she says, “When this bird died, scientists knew it was probably the last one.” At the time, cell lines generated from bird skin had just a 50 percent success rate—so Marlys asked them to send the specimen to two labs just to be safe. “They told me: ‘There is no other place.’” This lab was the only one with the capability at the time. As a technician, Marlys had developed a novel protocol to extract pulp from the tips of feathers, but the po‘ouli’s pinions proved too small. 

“I tried everything: skin, gonads, kidney, liver, spleen, heart—the cornea was too little—I couldn’t get it out, so I thought, Well, I’ll just take the whole front half and back half of the eye and see what happens.”

She checked her cultures constantly. Then one day, cells materialized. “The back groove of the eye was the only thing that grew,” she says. I can hear the relief in her voice almost twenty years later. 

 

Living cells from a northern white rhino. (San Diego Zoo Wildlife Alliance) 
 

When she offers a tour of the lab, I eagerly accept. “Don’t look up!” she calls to a colleague engrossed in pipetting solution as we enter. Sterile hoods and microscopes alternate along meticulous workbenches. Shapely bottles of pink liquid ready to be checked for signs of life rest on trays. They are surprisingly beautiful, like potions or perfume. 

“On any given day we have thirty to fifty cell lines incubating—mammals and birds take up to five weeks.” She opens an incubator and gingerly pulls out a tray. “Amphibians—up to a year. They seem to do better under hypoxic conditions, so we pump in a 2 percent oxygen gas.” After examining the bottles, she selects a likely looking culture.

“Have you ever seen a Chinese three-striped box turtle?” she asks. It’s one of the most endangered tortoises in the world and only found in Hong Kong. I shake my head and she slides it under the microscope. A brilliant constellation of actively dividing cells explodes across the computer screen. 

The image catches me off guard, like when the embryologist sprung on me a photograph of my daughter as a blastocyst right before my in vitro implantation procedure. I was in awe then, as I am now, knowing the fragile instructions needed to rescue a genetic line—spindles of sugars and phosphates and nitrogen bases—enfolded inside. Every turtle shell in the world receives its unique pattern from its parents’ genetics. This one has three dark stripes running the length of a spiraling, golden carapace. My daughter has my husband’s ears and his easygoing temperament, but when she belly laughs, her eyes crinkle just like mine.

“Care is pretty much around the clock,” Marlys comments, picking up a scrap of paper with what she calls “babysitting notes” for a Mang pit viper. Another colleague calls in sick, reminding the team to feed the cells by refreshing their nutritious growth medium and to keep an eye on temperature. 

“I’ve been doing this for thirty-seven years and it’s still fun—every day is repetitive, but different.” Her words echo anyone in the throes of nurturing new life. 

“When is your birthday?” she asks, reaching toward a row of scientific logbooks stacked above the workbench. At closer glance, I see that their brown-and-gold bindings are labeled in consecutive years. “This is my predecessor Arlene’s handwriting,” she says, flipping to January 1985. “On your birthday, we banked an oribi.” Neither of us know what that is, so we google it: a petite antelope with liquid brown eyes stares back. 

I ask her about the overcast February day we brought my daughter home from the hospital, wrapped in a blanket like a snuggly valentine. Marlys says that day brought six: a Guianan squirrel monkey, a Cooper’s hawk, a Guatemalan pit viper, a southern tamandua, a blue-crowned hanging parrot, and a Raggiana bird-of-paradise. The colors they invoke are dizzying. 

 

Baby sunflower sea stars, hatched using assisted reproduction and frozen sperm, stream across a petri dish trailing tiny suction cups. 
The recent success boosts new hope for the keystone species: in 2013 a mysterious wasting disease ran the coast from Baja to Alaska,
killing ninety-five percent of wild sea stars, rendering them functionally extinct. (San Diego Zoo Wildlife Alliance)
 

“A lot of the cell lines we store, like the Department of Fish and Wildlife’s condors, belong to others—we’re only caretakers,” she says, thoughtful. “In fifty years, no one’s going to know who I am.” 

She returns the logbook. 

“All they’re going to know is that we saved the cells. And that’s how it should be—just that they’re saved.” 

It’s time for her to check on her cell cultures again, and I need to get back to my family. Somewhere just beyond the Frozen Zoo, the new foal with a rake of spiky hair and soft little stripes on his legs nuzzles his mother.