Haploids, Bubble-Wrap and the Olympics

Over the past week or so, a lot has gone on here at Mahyco, mostly in the lab where as always we work sincerely but not seriously :)

This week I had the ultimate biological epiphany: not every organism has chromosomes in pairs of two. Now imagine me, sitting in front of someone that clearly knows what they are talking about, and blatantly saying, "No, that's not possible. That's not a thing."
And then Sheetal would sigh and flip over the sheet of paper, starting a third or fourth attempt at explaining what should have been an easy topic.
"Now...when you say 22 chromosomes in a set, you mean 22 pairs of chromosomes in a set."
"No, I mean 22 chromosomes make a set."
"Nahi! But chromosomes have to come in pairs or it won't even be a living thing!" I was getting really frustrated. I just assumed she was talking in theory, not in actual living things.
And this same argument carried on until finally, she said, "Only in humans! Humans have pairs of chromosomes, not plants!"
And then I had the epiphany of the decade, suddenly understanding the concept of polyploidy.
Thank you so much for putting up with me, Sheetal!


For another glimpse of what goes on here in the lab, imagine this:

In a multi-million dollar laboratory, up two flights of polished red stairs, hidden behind an elaborate security check-point system, past lines of computers, digital DNA freezers, turning right past the industry's most advanced laboratory robot, in front of two electrophoresis DNA chip-reading machines: you will find four apron-clad scientist popping a sheet of bubble wrap.

We have loads of fun here in the lab :)


The last highlight of my week is the Olympics. As I write this, we are watching the Indian archery team face off against the Japanese in the first round. It was a suspenseful match, and came down to a shoot-out after the 24-shot round was declared a tie. (won in the last stretch by the Japanese)
The coverage of the Olympics here is absolutely spectacular! They show every event, even the most unknown ones. I have found this type of coverage much more appealing, because you learn new games and find perfect strangers to cheer for until the end.
India has a good chance at gold this year in several events, and I can't wait to experience the excitement for them as the games continue.

My work here at Mahyco

Some quick updates:
We have nearly finished our initial project on Bacterial Leaf Blight in rice, and we are beginning the process once again with Sheath Blight. If you want to learn more about what I'm doing, just google search "molecular markers". Also, I sent my overview project to Ms. Fleming last week. For those of you reading this that don't specialize in molecular biology, here is that broad summary:

In the long run, we hope to identify those plants with multiple immunity QTLs so that the breeders can create a line of rice with lasting resistance to disease. It is much less likely that a pathogen can adapt to infect a plant with multiple resistance genes.

For my two month period in India, I am working in the Molecular Biology lab along side Sheetal Bhosle, my mentor and teacher. We are working to either validate previous studies on rice’s molecular markers, or to analyze genotypes based on pre-validated molecular markers. More specifically, we are focusing on analyzing Mahyco’s 34 rice genotypes for resistance genes to bacterial and sheath blight. Molecular analysis is necessary because to develop lasting resistance, a plant should have multiple different resistance genes. If a breeder were to attempt this the conventional way, there would be no way to differentiate between those plants with multiple resistance genes and those with only one.

A molecular marker is a piece of DNA sequence that is located near or on a gene that we wish to confirm in the genotype. For almost a century scientists have been tagging and arranging genes and gene-groups (known as Quantitative Trait Loci, or QTLs) that control favorable traits so that they can be identified on a molecular level, saving many generations of time for the breeder. The basic principle is this: the closer a marker is located to the gene you want within a chromosome, the higher the probability that one will not exist without the other. For example, a high-yielding plant can be tagged with two known genetic markers. In the next generation, if 1 in 100 plants have the first marker, but are no longer high yielding, while 10 of 100 plants contain the second marker but are not high yielding, we can conclude that the first marker is closer on the chromosome to the desired trait of high yield.

To begin our test across primers and genotypes, we first collected DNA samples from seeds. DNA extraction involves the crushing of the outer shell, disturbance of the cell wall, binding of the DNA to silica powder, then the use of water and ethanol to remove all polar and non polar contaminants.

Once the DNA is prepared, we can begin to test for the existence of our molecular markers that are linked with resistance to bacterial leaf or sheath blight. Each well in a 96-sample plate is given 5 micro liters of DNA. Then, a master mix including the forward and reverse primers, water a buffer, dNTP and Taq polymerase enzyme is added to each sample. The process that takes place in the PCR machine is thus: the samples are heated to denature the DNA. Next, the Taq enzyme replicated the process of DNA replication in our own cells as is facilitates the reforming of the sequence within our forward and reverse primers. In other words, as the system cools, the primers cut a segment of DNA which is then replicated. After 35 cycles, we should have 2 to the 35th copies of our desired sequence.

After the samples have completed their PCR amplification, we must test to ensure that the sequence was amplified before sending the samples to more precise equipment for analysis. To test for amplification, we place a row of randomly selected samples into an agarose gel. We place the gel in a buffer solution within a battery. DNA is negatively charged and will head one side as the cathode and anode create charged areas. Once completely diffused, the EtBr within the gel allows us to see the DNA under a black light. If it is clear that there are strands of more than 50 base pairs amplified, the PCR was successful.

For a more thorough analysis, the samples that are known to be amplified are placed in a MultiNA machine. This device works the same way as the agarose gel, but the computer can analyze the exact concentration of each specific base pair length within the sample. Though time consuming, we can use MultiNA results to confirm the existence of our desired marker and whether or not it was polymorphic.

The Vedic Expo

Here is the first in a series of short stories from my trip here in India...hope you enjoy! (Everything in these tidbits is 100% true story)

     As I watched the red sandstone and white marble spires appear over the horizon, I couldn't help but think: How many different famous temples can there possibly be in this city!? But, as with everything in Delhi, this one was nothing like I expected. We drove over a cobblestone hill, though the stones had been almost buried in sand due to lack of monsoon rains. As I gathered my things, I had a "conversation" with the driver- as I stepped out of the vehicle, I prayed that had understood my instructions. I would be back soon; this looked like a fairly simple landmark. One quick lap, some pictures, then finished and on my way to lunch…or so I predicted. I popped my scarf over my sweaty forehead and was off, kurti flopping in the dusty wind. After clambering about the temple to Krishna, I wondered why the driver had suggested several hours for this stop- it seemed entirely straightforward. I descended into the air-conditioned building below the temple, and attempted to blend in- needless to say, an American girl in Indian clothes, a headscarf and sunglasses receives the same number, if not more blank stares from others around her. I met the first glares with smiles, but after some time, I quietly made my way out. Clearly, tourists don't frequent this particular Hindu temple. On the way back up the slope, I noticed a neon poster with "Vedic Expo" splashed across in neat print. Elated at the chance to see some authentic Indian artwork, I veered into what would become one of the scariest, and later, most hilarious experiences of my trip. Ticket in hand, I was directed to the waiting area. In the distance, I could hear the guides arguing over whose English was best; he should be the for my tour. I could hear the whole conversation because, not only was I the only American at the entire monument…I was the only person at all inside the expo. Seconds later a man that looked to be in his forties, about my height and with an expression of indifference called me to a huge metal door: the start of the show. The door pushed sideways and disappeared into the wall to reveal nothing but shear blackness. I turned to my guide, wondering what the problem was. Had the power gone out? He responded by shining a flashlight into the emptiness. "Stand in the light," he directed.

Mouth agape, I must have appeared to be the stereotypical tourist of the decade. Stepping away, I stammered, "Why?"
He responded bluntly, "The show ma'am. Stand there, back to the wall."
Needless to say, I was not appeased by this answer. I stood there, helpless and confused. I was not about to enter a pitch-black room with a middle-aged man and stand with the door closed behind us. No-sir-ee-bob. My grandmother's voice rang in my head, "stay away from perverts…" (our family joke, which, in this case, took on a more serious tone) I looked, dumbfounded, back at the tour guide, and he burst into laughter. At his point, I am about to just leave in panic when I turn and notice the judgmental glares from the other employees. At this, I steeled my nerve and descended into the room, right where his flashlight indicated. Th guide shut the door and we were both engulfed in complete darkness.
To my surprise, he knelt down and pressed a button, bringing the whole room ablaze with light and color. I suddenly felt the weight of how stupid my fears were…although my expression in the sudden glow was enough to make the guide roar with laughter.
I breathed a sigh of relief as the lights danced in front of me, illuminating carved faces of krishna. The god's voice bounced off the walls and left no space untouched. I was annoyed to find that when the show was complete, the guide walked past me and opened a second door into darkness. Being relieved but still understandably anxious, I had already turned to escape from whence we came. The guide, however, motioned for me to follow, continuing deeper into what seemed like "Vedic spelunking".
Despite all its strangeness, the various rooms with their respective light shows were incredibly educational; topics ranged from the soul to karma to yoga and even reincarnation. The final stop was by far, the most "interesting": a metaphor for krishna's guiding hand within a "maze of confusion." Th tour had been moving along systematically until this final test against confusion…and my patience. 
Inside the final exhibit was a figure 8 of mirrors, complete with hexagonal-cylinders of mirrors filling in the loops. Inside these columns were two huge TV sets. The lights shown a dim gold and the screens displayed a simple PowerPoint slide of "Hare Hare Krishna Krishna". An inescapable loop of a men's choir singing these lines blasted throughout the room. I walked through the loop to find my guide, and I looked at him expectantly. He simply waited, appearing to me as if he wasn't going anywhere. After a few seconds of this staring stand-off, he signed for me to follow him round the figure 8. After some laps, we again reached the start, when he finally sighed and said, "You're supposed to become lost." I stood there, annoyed.
"I'm not lost. This is enough; I get the point."
I surveyed the small chamber and fixed my eyes longingly on the poorly-disguised door behind the guide. "There is no way out!" he said, following my gaze.
Frustrated, I ambled around once more, putting on the most ridiculous act of confusion: turning circles, seemingly awestruck at the complexity that was this bedroom-sized puzzle.
Pleased with my show, and himself apparently, he opened the door at last, washing us both in bright light from the lobby. He reached to shake my hand, but I pretended that I didn't see. Speeding out the door and leaving my water bottle behind, I never looked back. I met the driver at our pre-planned spot and we zipped away toward my next destination.


Delhi was a wonderful experience, but I just don't think my trip would have been complete without this strange event. So, I recommend that anyone in India see the Vedic expo...just make sure to bring a friend, and know what you're getting into!