you are saving me. 5 minutes ago I was crying from not being able to understand my molecular cell biology textbook (first year med student), and now it all made sense. thank you. love your videos, please never stop
i absolutely love your lectures! thank you so much for all the hard work you put in. you're saving our lives so we can become doctors and save many more lives!
THANKOU SO MUCH SIR! only if my lecturers could explain as awesome as you... I love your content and videos it ALWAYS helped me through understanding fully . i hope wherever you are you be always healthy and create more awesome videos. God bless you.
Hey, really good video. One question. In another video about cDNA you said that to separate the DNA/RNA hybrid we have to increase the pH, because at a basic pH RNA gets hydrolysed but DNA not, so we will remain with the sDNA, here you say we heat them up to separate, which makes sense for me, but which of the two methods are right? Or can we use both? Has one more advantage?
do we need to place our double stranded cDNA inside a vector of a prokaryote? (ie plasmid/lambda phage) or can we just insert the cDNA molecule directly inside a prokaryote??
Assuming you have determined the sequence of a certain enzyme/protein product, how will you identify its correct DNA sequence (*some codons are redundant or wobbled) using the cDNA libary? just asking.
For stuff that relies on alternative splicing (ex. antibodies) I think you'd only be able to make cDNA for one transcript variant that codes for one specific protein
Thank you sir, vividly explained but i have a little worry. is there a possibility for the sscDNA to be re-transcribe into RNA? And given that DNA polymerase needs a free 3' OH to be able to form the complementary strand or betterstill it needs a primer for elongation to take place. what happens in this case? where does the sscDNA get its primer from?
Out of curiosity I believe at 6.08 there maybe an error, unless I'm not thinking correctly. All eukar dna consists of polyA tails so therefore in order to prepare a cDNA library a poly-T primer is created for complementarity??
eukaryotic DNA doesn't have a poly A tail, it is the mRNA that does, when the reverse transcription is happening the poly A tail is removed and DNA does not contain poly T tail, therefore you don't need a Poly T primer in order to prepare a cDNA library
If restriction enzymes cleaves by recognition of some specific sequence.. Why does it selectivly cleave genes differently? Isnt there a high chance of braking middle of genes?
No, because then it wouldn't generate the same mRNA transcript due the 5'-3' directionality of DNA polymerase. The cDNA generated is an antisense strand (non-coding) which binds to the sense strand, or coding DNA.
wonderfully explained. I just have one problem with your lectures though. I have to have my inhaler next to me because you don't breath in between your explanations and that gives me short breath. It's not a joke I'm serious please take a breath sometimes. please!!
Amazing video, but I think there may have been some errors. Please correct me if I am wrong. 1. On the bottom right, the cDNA is shown to be 5' to 3' and the mRNA is also shown to be 5' to 3', should the cDNA actually be 3' to 5' so that is is complementary to the mRNA? 2. In 8:35, you say that you can take the double stranded processed DNA and place it in the eukaryotic bacterial cell, did you mean prokaryotic or am I confusing the meaning of your sentence and that the eukaryotic refers to the DNA?
For the first note, on the bottom right, the cDNA is shown to be 5' to 3' because it has been rotated 180 degrees. Notice the flat back bone is on top instead of the bottom of the bases, while the bases (perpendicular lines) are facing down. Imagine the cDNA was attached to the mRNA but then it was separated AND rotated 180 degrees to be put side by side so that they both are shown as 5' to 3'. I think that's what he did, but I am not sure.
yes actually the dna strand synthesised first must b 3'-5' nd cdna 5'-3' so thst it becomes complementary to the parent strand and has same polarity as the mrna strand...nd actually he meant prokaryotic bacteria
One application where cDNA is used is RNA-Seq: "While direct sequencing of RNA molecules is possible, most RNA-Seq experiments are carried out on instruments that sequence DNA molecules due to the technical maturity of commercial instruments designed for DNA-based sequencing. Therefore, cDNA library preparation from RNA is a required step for RNA-Seq." - RNA-Seq methods for transcriptome analysis. Hope that helps.
you are saving me. 5 minutes ago I was crying from not being able to understand my molecular cell biology textbook (first year med student), and now it all made sense. thank you. love your videos, please never stop
extremely clear and very easy to understand.
Just to be clear @7:16 did he mean "if we place this cDNA in a prokaryotic cell"
yeah, I'm also a little confusing at that point. I think it should be "cDNA in a prokaryotic cell"
YEs he made a mistake
Pannel Egboh lol, exactly, nonetheless his explanation is the best
Best Explaination 💯/💯, chow chow !
Yes. By which he meant vectors.
Thank you so much for making such a complex topic so simple and easy to understand. I wish my textbook explained things the way you do.
Thank you so much! Could not have understood this without your help.
Beautifully explained. Thanks very much for your great service.
Thank you... very easy to grab. Great teacher.
i absolutely love your lectures! thank you so much for all the hard work you put in. you're saving our lives so we can become doctors and save many more lives!
You make my life 10 times easier!! thank you thank you thank you❤️❤️❤️❤️
perfect!thank you
THANKOU SO MUCH SIR! only if my lecturers could explain as awesome as you... I love your content and videos it ALWAYS helped me through understanding fully . i hope wherever you are you be always healthy and create more awesome videos. God bless you.
really perfect explanation. your are one of the best lecturers
Thank you, you are awesome!!!
you are awesome man!!!!! what a great work
that was very well explained, thank you. could you please explain subtractive hybridization as well? :)
thank you
Hey, really good video. One question. In another video about cDNA you said that to separate the DNA/RNA hybrid we have to increase the pH, because at a basic pH RNA gets hydrolysed but DNA not, so we will remain with the sDNA, here you say we heat them up to separate, which makes sense for me, but which of the two methods are right? Or can we use both? Has one more advantage?
wonderful!!!
omg dis is very easy.....................l cant even believe this !!!! thank you very much Sir
wow 😍😍😍😍 super clear now!!
Very useful like other videos. Thanks.
Very much helpful
Sir have u posted lectures on plant physiology..I need them basically.
Very awesome no words to say
thank you i love you
Bravo!!!!
great lecture
Nicely explained.
Please make a video on Mitronchondrial Biogenesis
do we need to place our double stranded cDNA inside a vector of a prokaryote? (ie plasmid/lambda phage) or can we just insert the cDNA molecule directly inside a prokaryote??
Thanks
Is there some way to know each individual gene sequence
So that we can use a specific restriction enzymes to put the gene in a vector?
wow beautiful class
Assuming you have determined the sequence of a certain enzyme/protein product, how will you identify its correct DNA sequence (*some codons are redundant or wobbled) using the cDNA libary?
just asking.
Wow thanks man
great! but how do u prevent from alternative splicing when producing pre-mRNA into mRNA as a template?
For stuff that relies on alternative splicing (ex. antibodies) I think you'd only be able to make cDNA for one transcript variant that codes for one specific protein
Thank you sir, vividly explained but i have a little worry. is there a possibility for the sscDNA to be re-transcribe into RNA?
And given that DNA polymerase needs a free 3' OH to be able to form the complementary strand or betterstill it needs a primer for elongation to take place. what happens in this case? where does the sscDNA get its primer from?
nice class
clear and easy to understand, thanks man
thnx sir..
Out of curiosity I believe at 6.08 there maybe an error, unless I'm not thinking correctly. All eukar dna consists of polyA tails so therefore in order to prepare a cDNA library a poly-T primer is created for complementarity??
eukaryotic DNA doesn't have a poly A tail, it is the mRNA that does, when the reverse transcription is happening the poly A tail is removed and DNA does not contain poly T tail, therefore you don't need a Poly T primer in order to prepare a cDNA library
If restriction enzymes cleaves by recognition of some specific sequence.. Why does it selectivly cleave genes differently? Isnt there a high chance of braking middle of genes?
very easy now!
The mRNA strandis 5' to 3'. Shouldn't the complementary DNA strand be 5' to 3' also?
No, because then it wouldn't generate the same mRNA transcript due the 5'-3' directionality of DNA polymerase. The cDNA generated is an antisense strand (non-coding) which binds to the sense strand, or coding DNA.
Its easy to understand thank u..
Why it is called library !!?
So if I place a DNA inside a prokarytic cell, the prokaryotic cell cannot synthesize the protein, because of the introns sequences the DNA contain?
yes
wonderfully explained. I just have one problem with your lectures though. I have to have my inhaler next to me because you don't breath in between your explanations and that gives me short breath. It's not a joke I'm serious please take a breath sometimes. please!!
Wow
Amazing video, but I think there may have been some errors. Please correct me if I am wrong.
1. On the bottom right, the cDNA is shown to be 5' to 3' and the mRNA is also shown to be 5' to 3', should the cDNA actually be 3' to 5' so that is is complementary to the mRNA?
2. In 8:35, you say that you can take the double stranded processed DNA and place it in the eukaryotic bacterial cell, did you mean prokaryotic or am I confusing the meaning of your sentence and that the eukaryotic refers to the DNA?
yes i think youre right
I also think you're right
For the first note, on the bottom right, the cDNA is shown to be 5' to 3' because it has been rotated 180 degrees. Notice the flat back bone is on top instead of the bottom of the bases, while the bases (perpendicular lines) are facing down. Imagine the cDNA was attached to the mRNA but then it was separated AND rotated 180 degrees to be put side by side so that they both are shown as 5' to 3'.
I think that's what he did, but I am not sure.
yes actually the dna strand synthesised first must b 3'-5' nd cdna 5'-3' so thst it becomes complementary to the parent strand and has same polarity as the mrna strand...nd actually he meant prokaryotic bacteria
🤝
Please answer me ! I need help, why they are doing this cDNA, what are the domains of application
(sorry for my english, il french )
One application where cDNA is used is RNA-Seq:
"While direct sequencing of RNA molecules is possible, most RNA-Seq experiments are carried out on instruments that sequence DNA molecules due to the technical maturity of commercial instruments designed for DNA-based sequencing. Therefore, cDNA library preparation from RNA is a required step for RNA-Seq." - RNA-Seq methods for transcriptome analysis.
Hope that helps.
thanks
Using word eukaryotic for prokaryotic
An MNRA bacterial cells.
🥰🥰
1.5 speed... love you
same.
Sir the sound of ur videos r very low.
You probably might want to use speakers
I think labelling of intron and exon needs to be reversed.
I think I fell in love with you