Kaitlyn hunched over a styrofoam box on her bench. Nestled atop the densely packed layer of crushed ice inside sat strips of miniature plastic tubes tucked into a neon green rack. With practiced movements, she quickly transferred clear liquid from a tube in her hand into the strip tubes one after the other.
That’s the reaction mix aliquoted. She rolled her shoulders and straightened. Time for the template.
Reaching for a smaller pipette, she spun its dial until the display read one microliter. Then, carefully opening another set of tubes, she began transferring yet another clear liquid into the first.
At her elbow, Christa nodded in approval as she finished. “Good, good! You made sure not to cross-contaminate any of your reactions, from what I could tell. Now, you remember how to start the qPCR machine?
Kaitlyn nodded. She double-checked that her tubes were indeed closed and followed Christa to the machine in question. A rather futuristic white box with a touchscreen display sat next to the other various gadgets and gizmos that had become so familiar over the loops.
Placing the strip tubes inside the machine’s drawer, she quickly entered in a few parameters for number of cycles and cycle temperatures. She considered double-checking with her lab notebook, but was confident enough that it felt optional. Besides, Christa would tell her if she’d really messed up.
Instead, the woman nodded again. “Great! Wow, you really do know what you’re doing for an undergrad. Finished that whole thing without asking where to find a single thing. Have you worked in a lab before?”
Kaitlyn beamed at the praise. “Yeah, I have. One kind of like this, actually.”
“Really? Where at?”
“Um…” Kaitlyn hesitated, fishing for an appropriate lie. “It was an internship in high school. Anyway, thanks for the help. Guess I’ll just put the cDNA and stuff away and find you when it’s done running?”
“Sounds like a plan!” The woman elbowed her gently. “But to be honest, with how well you’ve handled yourself so far, I’m not sure you need me.”
Kaitlyn chuckled. “I’d still appreciate the sanity check. Just to see if the results make sense. And if they don’t, to figure out what dumb mistake I made.”
“Of course, of course.” Christa turned toward her own bench. “Well, you know where to find me!”
With that, Kaitlyn grabbed her ice box and headed toward the freezer. The interior of the thing was practically bursting full of boxes, bottles, tube racks, and other assorted bits and bobs from around the lab. How anyone managed to find things in here never ceased to amaze her.
Heck, I don’t know how I find things in here either. Reaching inside, she retrieved box after box from the iced-over shelves and placed her reagents back inside. Must be those memory tricks coming in handy.
With that done, Kaitlyn heaved a sigh and headed toward the clean hood. Her qPCR reaction would be running for hours yet. Might as well take care of some plates in the meantime.
Throughout the loops, she’d watched and performed this assay a number of times. On its own, it was pretty straightforward. But putting it all together as a full experiment felt like a new challenge all its own. From growing the plants, treating them, harvesting the tissue, extracting their RNA, converting it to DNA, and now finally quantifying that DNA with qPCR? It was pretty involved. The whole thing made her especially thankful for Christa’s watchful eye.
This just makes me realize how hard it’s gonna be to optimize experiments. Kaitlyn grabbed a few sterile dishes from underneath a cabinet. Things can go wrong at pretty much any part of that process. It’ll save me so much time if I can get them to work reliably though. It’s gotta just be a matter of figuring out the right variables.
But she felt that was all the more reason to go for it. Having to redo an experiment because she messed it up was one thing. But knowing in advance the most efficient way to prove or disprove a hypothesis? Surely that was valuable. It would save so much time, not having to slog through iteration after iteration of tweaks to find the best assay design. Especially given the short timeline of each loop.
First thing’s first - I need to get the technique down. Then I can focus on optimizing the treatments and RNA extraction. The clean hood turned on with a familiar whoosh as Kaitlyn opened it. She let it run for a bit while collecting her other materials. I remember that Freddy complained about the extraction part a few times… Maybe I can ask what he does and compare. He could do something different than Christa.
Settling in, she smiled. First thing’s first, though. Get qPCR down. Don’t screw it up. Then, in a few loops, it’ll be just as second nature as everything else.
Ensure your favorite authors get the support they deserve. Read this novel on Royal Road.
***
Four weeks of repeatedly running qPCR later, Kaitlyn was convinced that she was indeed screwing this up.
How. HOW. Her teeth gritted in frustration as she glared at the graphs before her. How is there STILL signal there?!
The source of Kaitlyn’s anger stared dispassionately back through the display. A collection of curves that quantified her gene of interest clustered together like a handful of snakes. Those weren’t the problem though. No, the problem was what wasn’t there.
We put in a negative control for a reason. There should be at least three graphs that are just boring, flat lines. Frantic taps of her fingers cycled through the graphs until the controls in question highlighted. So why aren’t they flat?! How is there signal?!
Christa hummed thoughtfully as she peered over Kaitlyn’s shoulder. “Well, that’s not good. Same thing as last time?”
“Yes,” she sighed with exasperation. “I don’t know what I’m doing wrong. I made sure to switch tips every time and kept the controls closed unless I was working on them. I even got new nuclease free water!”
“Hmmmm…” Christa patted her shoulder comfortingly. “We’ll figure this out. Don’t worry. In the meantime though, it’s looking like we can’t use this data though. Sorry, Kate.”
Kaitlyn slumped in her chair. “I know. It’s just… Is there something you see I should be doing? Is it me?”
“I honestly couldn’t tell you.” The woman frowned thoughtfully. “Have you been wiping down your bench? Maybe there’s a contaminant there.”
“Every time. I just got my lab coat cleaned, too. Freddy even let me borrow his exonuclease spray.”
“Really?” Her frown deepened. “Hmmmm… I don’t know then. Here, next time maybe we both prep reactions and see if they both fail. How about that?”
Kaitlyn nodded dumbly. At least that’ll tell me if it’s really my fault. That or some cosmic joke. This is so friggin’ frustrating…
“What’s going on over here?” Elliot peeked around the corner and glanced at the screen. “qPCR?”
“Yeah. Kaitlyn’s been having trouble with her controls getting contaminated.”
“I swear I’ve tried everything I can think of…” she lamented.
Elliot whistled. “Wow. Have you tried an exorcism? Your bench might just be cursed, you know.”
A withering glare accomplished nothing but making the guy smile. “Just saying. Don’t knock it till you’ve tried it.”
“We’ll repeat the assay again next week,” Christa reassured her. “For now, I think you need some rest. You’ve been running yourself ragged over this thing.”
With a final comforting pat, the short woman headed back toward her bench. Kaitlyn just laid her head in her arms. I can’t even get the technique right here. That’s step one! Maybe I started on something too complicated? It’s not like I’m trying to do a Western blot or something really fancy, here. This is supposed to be pretty straightforward. So what am I doing wrong…?
“Don’t beat yourself up about it too bad.” Elliot’s voice made her lift her head. He leaned back against the counter at her elbow, casual as ever. “Stuff happens. Even to ‘real’ scientists. You should’ve seen Freddy when he started out with his pollen assays...”
A frown creased her brow. “What happened?”
“The guy went through the methods of a paper and tried to reproduce them. Not even changing anything, he just wanted to make sure the assay worked.” The guy shook his head with a smile. “Took him about four months. Eventually, he had to call the original authors and get them to walk him through step by step. Apparently that did the trick, even if he swears up and down they had him doing the same exact thing. Sometimes an assay really just doesn’t agree with you..”
Kaitlyn groaned. “That’s well and good if this were some special new assay. But it’s just qPCR! Everyone does it. It shouldn’t be that hard to get right.”
“I guess, but still.” Elliot shrugged. “Getting all frustrated certainly won’t help much, will it?”
“No, but I am frustrated.” A scowl crept onto her face. “And I will be until I figure out what’s going on.”
The ginger grinned. “Now you sound like a real scientist. Stubborn as a mule.” He pushed off from the counter. “Well, I gotta get back to it. Good luck figuring things out.”
Kaitlyn watched as her fellow undergrad headed toward the growth chamber. Along the way, he stopped for a quick chat with Esme. The raven-haired girl’s tinkling laugh echoed down the hall to reach her ears.
Maybe they’re right. Maybe things do just happen sometimes. But… another sigh escaped her. I could accept that once or twice. But three times in a row? There has to be some deeper reason here. Some real underlying cause.
She already knew that small things like this were prone to minor inconsistencies - Scarra’s tiramisu gifts were a clear example of that, not to mention the other bevy of experiments that suffered from variations. But this was different. This felt just like the assay was taunting her. Laughing at her.
Kaitlyn frowned at the incriminating graphs once more. I’ve gotten a lot better at hunkering down and focusing on things. At putting in real work. I thought that acing my classes was the real test of that, but maybe I was wrong. Maybe… it was all working up to this.
She pushed back from the counter and stood. Right. I wanted to focus on getting better at lab, right? Time to get serious about it. I’m not gonna rest until I’ve beat this damn assay.
***
Cycle 24 Complete!
Ending: Under the Radar
Total Endings Unlocked: 6