I Am Diagnosed as a Medical Titan

Chapter 175: Chapter 134: Sequencing Before the Dawn (Part 2)

”Huh?” Cheng Xiyao paused. ”Your sister-in-law is here?”

”That’s not the point!” Yi Xiangwan said nervously. ”Is this really an emergency? But Boss, we’re all rookies. Why didn’t you get the people from the clinical lab at Affiliated Hospital No. 1?”

The answer was simple.

The senior staff in the clinical lab had their own established procedures.

If Jiang He tried to direct them, they would question why he wasn’t adding this reagent or why he was changing that parameter.

The communication overhead would be too high.

But these six were different.

Over the past several weeks, Jiang He had put them through a hellish training regimen on the theory of RNA extraction.

They followed his every word, and they would execute his procedures without the slightest deviation.

So, Jiang He said, ”Listen up. The hospital has admitted a patient with an extremely high-risk case of severe pneumonia from an unknown cause. I suspect it’s a new, recombinant flu virus. Tonight, we need to extract the viral RNA, reverse transcribe it into cDNA, run PCR to get amplified fragments, and then put it on the sequencer to obtain its genetic sequence for confirmation.”

The six of them: ”???”

Tang Pei: ”Bo-Boss... this unknown virus... is it... safe for us to work on?”

”The sample we received has been thoroughly lysed and inactivated with guanidinium thiocyanate. It’s just a pile of nucleic acids now; it’s not infectious. The underlying principle is identical to the cell-free miRNA extraction from peripheral blood we trained on a few days ago. In fact, since we’re extracting the viral genomic RNA, the abundance is much higher, which actually makes the procedure easier.”

”The key is to prevent RNases in the environment from degrading the sample.”

Jiang He said sternly, ”I trained you all myself. I have absolute confidence in your lab discipline. For tonight’s procedure, every single step must be performed exactly according to the protocols I taught you. No extra movements, no deviations. Understood?”

”Understood!” Lu Xiaolin, the oldest of the group, was the first to respond.

”Got it, Old Jiang. Just tell us what to do,” Chen Hao said, slapping his cheeks to wake himself up fully.

”Go to the changing room. Double isolation gowns, double latex gloves, N95 masks, and goggles. Then enter the core area.”

「3:15 a.m.」

Ke Zheng arrived at the lab entrance carrying a sealed biosafety transport container.

He handed the container to Jiang He. ”Jiang He, this is Mark’s deep bronchoalveolar lavage fluid. It’s already been inactivated with lysis buffer.”

”Got it.”

Jiang He took the container, turned, and swiped his card to enter the lab’s core area. He immediately began giving orders:

”Group One, Chen Hao, Xiangwan, you’re on reagents. Open fresh bottles of everything—DEPC-treated water, absolute ethanol, isopropanol. Make sure there’s absolutely no RNase contamination. Spray and wipe down the workbench twice with RNaseZap.”

”Group Two, Cheng Xiyao, Tang Pei, you’re responsible for fitting all the pipettes with filtered tips. Get the centrifuge tubes and ice boxes ready.”

”Xiaolin, Yizhou, you two assist me. Get ready for the column extraction.”

Standing before the laminar flow hood, Jiang He opened the transport container and took out the cryotube containing the inactivated sample.

”Let’s begin.”

Inside the lab, the atmosphere was serious and efficient. This group of young students was now demonstrating astonishing discipline.

’This is the result of their training!’

Jiang He picked up a P1000 pipette and fitted it with a filtered tip.

”Add one milliliter of Trizol reagent.”

He transferred the sample into a new centrifuge tube and added the Trizol.

The clear liquids mixed instantly.

”Let it stand at room temperature for five minutes to allow the nucleoprotein complexes to completely dissociate.”

Jiang He glanced at the wall clock.

Once five minutes were up, he said, ”Add 0.2 milliliters of chloroform.”

”Cap it tightly. Vortex vigorously for fifteen seconds.”

”Let it stand for three minutes.”

Beside him, Gu Yizhou had already opened the lid of the high-speed refrigerated centrifuge, which was pre-chilled to 4°C.

”Place it in the centrifuge, balance it, 12,000 rpm for fifteen minutes.”

Those fifteen minutes felt incredibly long.

”BEEP—”

The centrifuge stopped.

Jiang He carefully removed the centrifuge tube.

Under the cool light, the liquid inside the tube had clearly separated into three layers:

At the bottom was the red phenol-chloroform phase, in the middle was a thin, white protein interface, and on top was the colorless aqueous phase.

All of the viral RNA was dissolved in that top, colorless aqueous phase.

Jiang He took a deep breath and switched to a P200 pipette.

This was the most critical step in the entire extraction process.

If he aspirated any of the middle protein layer or the bottom organic phase, the entire sample would be ruined.

He braced his elbow on the bench and pressed the pipette’s plunger.

The tip hovered precisely in the center of the colorless aqueous phase as he slowly drew up 400 microliters of liquid, transferring it to a brand-new, nuclease-free centrifuge tube.

Throughout the process, Jiang He’s hand didn’t tremble in the slightest.

The steady hand of a surgeon was on full display in that moment.

Lu Xiaolin watched from the side, his scalp tingling in awe.

’Even veteran researchers with years of lab experience would have a slight tremor when aspirating the aqueous phase free-hand, but Jiang He’s hand was as if it were welded in place—terrifyingly steady.’

”Add an equal volume of isopropanol. Chen Hao, hand me the ice box.”

After the sample sat on ice for ten minutes, it was placed back into the refrigerated centrifuge.

12,000 rpm, ten minutes.

When it was removed again, a tiny, translucent, gel-like pellet had appeared at the bottom of the tube.

”The RNA has precipitated,” Cheng Xiyao said, her eyes lighting up behind her goggles.

Jiang He: ”Discard the supernatant. Add one milliliter of 75% ice-cold ethanol to wash the pellet.”

After gently resuspending the pellet, he centrifuged it for a third time.

7,500 rpm, five minutes.

Finally, he used a pipette to very carefully aspirate all of the ethanol, then placed the open centrifuge tube in the airflow of the clean bench.

”Air-dry for five minutes. Don’t let it dry out completely, or it will be difficult to dissolve.”