Thank you very much! I was just not getting the principles of micro-array, but this video clarified everything in less then 2 minutes! Once again thank you!
I use Trizol reagent to purify, which is similar to a phenol/chloroform precipitation. You just have to make sure that you eliminate genomic DNA, and DNAse digestion on RNeasy columns from Qiagen can do that. Then you can do the reverse transcription to make the cDNA. Also, for RT-PCR, primers should be made to span an intron so that you can eliminate any possible signal from genomic DNA contamination.
You would typically have two different batches of cells, grown separately in culture. One batch of cells would then be infected, and after a period of time, both batches would have their RNA isolated and reverse transcribed and used for subsequent analysis.
Micro arrays are so cool! But remember that it measures RNA expression not necessarily protein expression! The effects RNA interference, RNA binding proteins, RNA degradation/ turnover, can skew this kind of data.
befz88 before they mix them up they flourescently label the sample (red for infected, green for uninfected). After this step, then they mix them up. so thats how they differentiate.
2 tissues and each tissue fluorescent coloured differently. The damaged tissue and the healthy tissue can either act the same or differently. If it acts differently, the DNA sequence will be different, and thus not bind to certain spots where the healthy DNA can bind. Because they are fluorescent labelled, the laser can pick up the color wavelengths in the spectrum, red (Cy5, 670nm) and cy3 (green 570nm). So the colour will tell how the healthy tissue is expressed compared to the damaged tissue.
Hey great video. I have a question... when the fluorescently labelled cDNA binds onto the microchip, what is actually in the microchip. Is it the DNA taken from different bacterial strains? For instance is the DNA for one bacteria extracted and put into one of the spots? Thankyou in advance
Each spot(hole) in the chip contains different oligonucleotides(about 20 nucleotides long) complementary to unique regions of the genes you are investigating, so that they bind while other genes are washed away. hope this helps
Question: Why combine the samples from the infected and uninfected cells together? Surely you'd want to *compare* what's being expressed by keeping the samples separate and reading them side-by-side, rather than all of it in one chip?
actually it makes sense, if there are same strands of dna on both green and red samples then yellow color will appear from the combination of the two colors, if the strand is just on one of the samples either green or red will appear, the rest will be empty. It seems easier to compare that way
+promise no , analyze gene expression. They are machines able of making several hybridization experiments. learn.genetics.utah.edu/content/labs/microarray/
@IsaacDLovesChrist Make cDNA first. Then your primer set you can either design it your self or pay a company I recommend paying save some man hours and get it done with QC.
This is the best video out there on DNA microarrays and it goes for like 1:30.
Glad you like it!
Thank you very much! I was just not getting the principles of micro-array, but this video clarified everything in less then 2 minutes! Once again thank you!
I'm glad this helped you!
@guydori.. What do you study
This is the type of video that makes me come to youtube for fundamental understanding of a concept.
this was the best videos I've seen so far. Very short and to the point. Thanks
Thank you for the teaching of micro array of DNA.
Bless you.
Wow the video is helpful even after 14 years! Thanks alot
Great Video! We need more of this...to help understand science! Thank you!
thank you so much for such a great infrormation ,this video clarrified the principal of microarry in such an amazing way
thanks!!! u helped me understand microarrays right before my final
Thanks for the detailed illustration 💙
So much better than my professor's explanation, and in just 1.5 minutes!
16 years ago,damn. saved me before midterm
I like the background song, really relaxing.
such a great video! thank you
wow, your very knowledgeable. thanks for all your help.
Thanks a lot, you guys are great.
I use Trizol reagent to purify, which is similar to a phenol/chloroform precipitation. You just have to make sure that you eliminate genomic DNA, and DNAse digestion on RNeasy columns from Qiagen can do that. Then you can do the reverse transcription to make the cDNA.
Also, for RT-PCR, primers should be made to span an intron so that you can eliminate any possible signal from genomic DNA contamination.
great explanation, Thanks!
Best test to detect irregularities in environment.. Thanks
perfect.. thank you its very help full
nicely explained. well done
Freeking awesome!
Beautifully explained! Thanks
very helpful video!
nicely done !!!!!
This is so cool omg
Loved it
thanks for your helpful video
Very good. Thank you.
Medicine student, Triest, Italy.
Hope things are going great for you
You would typically have two different batches of cells, grown separately in culture. One batch of cells would then be infected, and after a period of time, both batches would have their RNA isolated and reverse transcribed and used for subsequent analysis.
@Proneural What is your process for designing the primers? Which company do you use to generate your RNA primers?
Nyc animation...helpful stuff indeed!:)
what a fascinating technology
@arepeace0687 dont the complimentary strands in total just form the whole genome of the organism? from a gene library that was available?
Thank you
good one, thanx
Micro arrays are so cool! But remember that it measures RNA expression not necessarily protein expression! The effects RNA interference, RNA binding proteins, RNA degradation/ turnover, can skew this kind of data.
Is there concurention between the infected and non-infected DNA pieces on these wells?
Are you sure the CDNA is double stranded? perhaps it could be heated/melted before applying to the microchip?
I'm glad that 600 000 peoples see this video
Me too! :)
how do you know which one is infected and which wasnt if you mixed them before the array?
awesome
Perfect
ooohhh now i get it! cool stuff :D thanks for uploading the vid!
where are you now after 11 years
befz88 before they mix them up they flourescently label the sample (red for infected, green for uninfected). After this step, then they mix them up. so thats how they differentiate.
This is the shortest and the simplest video on microarray for identification of plant pathogens
2 tissues and each tissue fluorescent coloured differently. The damaged tissue and the healthy tissue can either act the same or differently. If it acts differently, the DNA sequence will be different, and thus not bind to certain spots where the healthy DNA can bind. Because they are fluorescent labelled, the laser can pick up the color wavelengths in the spectrum, red (Cy5, 670nm) and cy3 (green 570nm). So the colour will tell how the healthy tissue is expressed compared to the damaged tissue.
I WAS THE 1000000TH VIEWERRR
@Proneural Good choice!
Hey great video. I have a question... when the fluorescently labelled cDNA binds onto the microchip, what is actually in the microchip. Is it the DNA taken from different bacterial strains? For instance is the DNA for one bacteria extracted and put into one of the spots?
Thankyou in advance
I agree. what is it?
Each spot(hole) in the chip contains different oligonucleotides(about 20 nucleotides long) complementary to unique regions of the genes you are investigating, so that they bind while other genes are washed away. hope this helps
Superb
at 0:37 where did the complementary DNA come from? how did they stick there?
แปลไทยให้หน่อยได้ไหมอ่ะค่ะ
แล้วในmicroarray ใส่probe หรือ Dna เราไม่เข้าใจ
And to isolate the RNA you would use phenol chloroform extraction? and then make cDNA from the RNA you collected?
DAD IS THAT YOU ?
+Cameron Steele Yea
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thanks dog love you
1:02 LMAO THATS MY OLD PI's BOSS JIM CARRINGTON!!!!!
Thanks for only 16 subscribers 😢
i still cant get it
How are they fluorescently labeled?!
i still don't understand how expression level can be calculated :(
Who else came here because of RP's RJ?
hello adam
I love u.
Question: Why combine the samples from the infected and uninfected cells together? Surely you'd want to *compare* what's being expressed by keeping the samples separate and reading them side-by-side, rather than all of it in one chip?
my thoughts exactly
actually it makes sense, if there are same strands of dna on both green and red samples then yellow color will appear from the combination of the two colors, if the strand is just on one of the samples either green or red will appear, the rest will be empty. It seems easier to compare that way
Mondestrasz
Yeah, I'd actually made sense of it after a little thought - probably like 5-10 minutes after I posted it. :P
Mondestrasz Precisely. *nice Dante icon, btw!*
cDNA is double-stranded, how can it binds to mRNA?
please i dont know the purpose of microarray ?
explaine in details pleas
+promise no , analyze gene expression. They are machines able of making several hybridization experiments.
learn.genetics.utah.edu/content/labs/microarray/
@IsaacDLovesChrist Make cDNA first. Then your primer set you can either design it your self or pay a company I recommend paying save some man hours and get it done with QC.
Why is cDNA more stable than mRNA? (0:12)
@befz88 Ah by labeling them very carefully! Who wants to redo do experiments because of mislabeling!?!
Absolute expression levels cannot be calculated from a microarray. Only relative expression, that is the ratio of red to green.
probably because it was a stupid question and obvious which i now notice after growing up a bit haha
To electonecly what??? at 1:10
electronically capture the data
i was just kidding
how can it be any easier.
thanks anyway
2019?
is morgan freeman narrating this? o.0
No they can be. When you do single-channel array
Dude, that is so elementary. How can you not know it?
Microarrays are confusing.
peng
not clear enough
this sucks