1) Next Generation Sequencing (NGS) - An Introduction
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- čas přidán 21. 06. 2015
- What is Next Generation Sequencing?
➜ Next Generation Sequencing (NGS) is a powerful platform that has enabled the sequencing of thousands to millions of DNA molecules simultaneously. This powerful tool is revolutionizing fields such as personalized medicine, genetic diseases, and clinical diagnostics by offering a high throughput option with the capability to sequence multiple individuals at the same time.
Corrections:
➜ [0:35] Sanger Sequencing was developed in 1977, not 1900s
➜ [2:22] The video refers to an "Illumina MySeq" machine - the correct name of the machine is actually "Illumina MiSeq"
For more information on Next Generation Sequencing (NGS), please visit:
➜ Knowledge Base: info.abmgood.com/next-generat...
For a list of other free NGS resources, visit:
➜ info.abmgood.com/next-generat...
Thank you to all of the amazing community contributions towards translating this video into other languages including:
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(To view the translations, toggle the CC button and then go to Settings ➜ Subtitles/CC to select a language.)
Watch the other videos in this series on NGS:
➜ Sample Preparation: • 2) Next Generation Seq...
➜ Coverage & Sample Quality Control: • 3) Next Generation Seq...
➜ NGS Playlist: • 1) Next Generation Seq...
Check out our other video series:
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➜ Adeno Associated Virus: • 1) Adeno Associated Vi...
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Here's my homework summary:
0:06 General overview
0:12 What is NGS?
0:33 Sanger sequencing
2:16 NGS overview
1) 2:39 Sample preparation
2) 2:51 Sequencing machines
3) 3:11 Data output
3:23 Different NGS platforms
1) 3:40 Pyrosequencing
2) 4:33 Sequencing by synthesis
3) 5:31 Sequencing by ligation
4) 6:42 Ion semiconductor sequencing
7:19 Comparison of NGS platforms: coverage of genome per read
8:26 Applied Biological Material's NGS services
I must say, this video is probably more helpful for any Bs, Ms or PhD student than a 1h lecture by some Professor...
High educational value!
Thanks, ABM!
Thanks for leaving the timestamps for all the sections! I'm sure others will find it useful. Also, thank you for your very nice comment. We're happy to hear that you found the video helpful!
Now you don't have time to do your homework!
@@bogdanbogdanovich140 "Be not afraid of going slowly, be afraid only of standing still" - Chinese proverb ;)
agreed!
Be of great help to write my report.
A very comprehensive video for anyone who wants to get a brief overview on NGS, the methods and the comparison between them under 10 mins. Highly recommended.
Hello Shekar,
We are really happy to hear that you found the video helpful ;) Please check out our other videos too!
I have been trying to understand the specifics of NGS for days now, and I understood it all after one watch. Thank you so much
Hi Antonin, thank you for watching our video and leaving such a nice comment! Please let us know if you have any questions about the material and we'd be glad to help clarify!
we have a few of your pcr machines just for the ramp speeds. my 6-9kb kb assays always have great bands. keep it up!
This was an excellent video! I enjoyed it. I think it is worth mentioning that the helicos sequencer, one of the NGS technology, does not require amplification during the sample preparation time.
Explains in 5 minutes what my lecturer tried to in 30 mins....straightforward, uncomplicated video.
However like similar comments read, the woman talks far too quickly and the background music is an unwanted distraction.
Hi Timothy, thanks for your comment! This was one of our earlier videos, so we've been working on improving the audio in our newer releases based on everyone's feedback :)
This is a really good brief summary of the most commonly use NGSs. I watched that the night before my exam and had them all stuck in my mind
We are glad to hear the video was helpful. Thanks for watching!
I'm glad this exists. This was the only simple explanation of NGS I could find.
Thanks for your comment, Harrison! We're happy to hear that our video is helpful :)
Great video, good explanations and perfect speaking speed! Literally got me a first class honours on a 'Compare & Contrast the NGS Platforms' final year assignment! Appreciated :)
this was very helpful & informative. I liked how the video notes were typed out so, instead of frantically taking notes on what was said, I could focus on the process and animations. Also, the overall diagram on the page is amazing.
+Samantha Seager Hi Samantha, thank you for your comment! We're glad you enjoyed the video!
Where are the video notes? I don't see them anywhere, but that would be very helpful.
Hi abm, thank you for an really great video which clearly explained the differences between these platforms. It was very helpful to have the text and a clear image of the steps involved. Loved the animations.
Thank you for your nice comment :) Happy to hear you enjoyed the video.
!546?!
I'm a french medical student and you've helped me a lot understanding biochemistry ! thank you !
Hi Arnaud! Thank you for your comment and we're glad our videos helped. FYI, we do also have a comprehensive knowledge base on the topic of NGS that might also be useful to you: goo.gl/Ce0M4O
Dear Arnaud!
I need some useful Lecture Notes/Books for Biochemistry and Molecular biology.If you,ve then please send me on my email address:asif.phw66@gmail.com.Thank you
Interested in Next Generation Sequencing? Learn more by watching our new introductory video on NGS - czcams.com/video/jFCD8Q6qSTM/video.html
+Applied Biological Materials - abm The videos are great but Could you decrease the background music. It is difficult to follow the speaker.
Thanks for your comment, +beagarwa, we will take note of that in future videos. In the meantime, we've done close-captioning for all our videos -- hope this can help.
Great video! However, i have some doubts regarding sequencing by synthesis, as i haven’t really understood the process, are there any videos i can find specifically on this topic?
Best video on this subject that I've seen so far
Hi Jan, glad you enjoyed our video! Please let us know if you have any questions :)
NGS seems like a useful tool for the home kitchen. For example, if I want to know if my italian extra virgin olive oil really contains italian olives. As a student I face this problem every day, since I can't afford expensive italian brands.
Nice video..Thanks to 4k Video!!
Very clearly analysed video... Thank you for your brief description.
No problem! Glad you found the video helpful ;)
Thank you for your information, I would like to know the NGS (4.6) it’s ok? Kindly please your help to clarify?
Thank you in advance.
Which book is good to study next generation sequencing
great overview of NGS technologies!
Glad you enjoyed it, Brenda!
Easy to understand. Helped a lot. Can We get these slides?
Sequencing by synthesis is used by Illumina. It is step-by-step incorporation of a reversible fluorescent and a terminated nucleotide. All the bases are added to the sequencing chip. Once a base is incorporated, the rest are washed away. The fluorescent is read and recorded. The fluorescent marker and the terminated nucleotide is removed. This process is repeated until gene sequencing is complete. It eliminates the homopolymer error issue but it was has increased error due to increased read lengths.
Hello Seeba,
Thank you so much for your input! It definitely helps to make our video more informative.
Stay tuned for upcoming videos too!
Amazing explanation. Thank you
Hi Terrence, thanks for your comment! Glad you enjoyed our video -- if you would like more info about NGS, feel free to visit our knowledge base: goo.gl/Ce0M4O
Why is sequencing by ligation offset by one nucleotide?
How many kinds jobs are available in market aspects of Next generation sequencing right now ?
very nice general introduction for people who are not experts in the field
Thanks for your comment! We're happy to hear that :)
helpful video for biginner level
very explanatory, thank you
Hi ABM, do you have a video about the detailed process on sequencing by synthesis applied by Illumina High Seq? Thanks
Hi Angelbert, thanks for watching out video and leaving a comment! We have more information over at our knowledge base (goo.gl/Ce0M4O) -- most of our knowledge bases and videos are based on the sequencing by synthesis method. Did you have a specific question in mind?
I am suffering from schizophrenia. i am male going for Pre Implantaion Genetic Test for the disease. I contacted Medgenome Genetic Lab and came to know that PGT is possible for Schizophrenia. Is PGT possible for Schizophrenia? Please reply me
thankyou so much your explaination is awesome it clears my doubt but will you please elaborate more about sequencing by ligation method
Hello there,
We are really glad to hear that you found the video helpful! :) and of course we will be happy to assist you further. Please contact us at technical@abmgood.com
Amazing video thanks.. please reduce the speed of audio.
I wish you did not use the background music.
I LIKED IT!
I wish the same
It's not so annoying :(
What is read and unread??
Can I get your videa's script?
I've really enjoyed the video, keep it up
Hi Asizu, thanks for watching and glad you enjoyed our video! Let us know if you have any questions!
I like this video, it helped explain something initially confusing for me
We're happy to hear that, Debbie :) Thanks for watching!
Excellent video and very explanatory, thank you!
Thanks! Please let us know if you have any questions, we'd be glad to help!
well, a few more details would be fine, like what are the terminator gaps in sequencing by synthesis and how they are cleaved!
Hi DUFo476! In general, Sequencing by Synthesis (SBS) technology uses fluorescently labeled dNTPs which act as "reversible terminators" for polymerization. During each sequencing cycle, a single labeled dNTP is added to the nucleic acid chain. Then the fluorescent dye is identified using laser excitation and imaging. Finally, the reversible terminator is enzymatically cleaved to allow for the next round of dNTP incorporation to occur.
Does this answer your question? If not, please elaborate on your question and we will provide you with more information!
Great video! Although the music is a little distracting
Hi!
Of course, it is much easier to perform a microarray analysis on gene code but a miseq analysis provide a higher significance of result. Though its expensive but it is usually worth it. I would like to know what are all the advantages of using Miseq over microarray?
Many thanks!
Hi Sebastian, Microarray relies on hybridization of DNA fragments to probes on a chip. To synthesize probes for microarray, you will need to know the sequence ahead of time. Thus, microarray can only examine known sequences either for copy number or gene expression. Sequencing is an unbiased approach to examine all the sequences, known and unknown.
For studies that require quantification, sequencing can certainly provide better quantification. E.g. how many reads aligned to this gene or this DNA region has X times coverage.
Of course, sequencing can provide base-pair resolution on genomic variations and breakpoints while microarrays (such as aCGH) can only provide a rough estimate where the breakpoint may be.
Hey the knowledge base link isn't working. Do you have a working link?
Hello Pixl,
It seems like the link is working on my end. Please copy and paste this URL on your browser and see if it would work: info.abmgood.com/next-generation-sequencing-ngs-introduction
Nice video, but I can't seem to open the link to your knowledge base... Is there a site maintenance or something?
Hi Svetoslav, I have just checked the link and it seems to be working! Perhaps there was momentary site maintenance. Thank you for pointing this out!
very nice..
thanks for video which helped me a lot...
Hi Mushtaq, thank you for your comment -- glad you enjoyed our video. Be sure to check out our knowledge base, also: goo.gl/Ce0M4O
Very informative. Thanks.
Hi Thenmoly, thank you for your comment! Glad you enjoyed our video!
I am interested in Scenario based Questions on NGS as I am preparing for my terminal exam in Molecular biology. Thank you
Hi Asif, sorry, we don't have any scenario-based questions on NGS available! We do have knowledge base articles on NGS if you'd like written materials to study from: www.abmgood.com/marketing/knowledge_base.php
Hi, minute 1:53 how are you sure that the there is a a flourescent ddnucleotide at each location? Isnt there a possibility that at least one location a flourescent ddnucleotide did not pair? Thanks for any clarification.
Hi Muhammet, thanks for watching and commenting! Normally, sequencing experiments are performed several times in order to have confidence in the accuracy of the sequencing results. This term is called "coverage" (or "Depth) -- you can read more about it on our knowledge base: goo.gl/o3gjDe
What can be a 'coverage of genome per run' up to now ? Thanks
Hi Ashok! Thanks for watching and for leaving a comment. The coverage of genome depends on the genome size and the sequencing machine. Using a typical HiSeq platform today can achieve about 30X coverage for a human genome per lane. Hope this helps!
Thank you for this video
Your very welcome, Allen! Please let us know if you have any questions regarding the material!
This video was AMAZING
Thanks!! :D
Hii.. M rasmi. This topic is very interesting. And my seminar topic is also NGS. So can u please help me about this topic??
How reliable are the sequences of whole virus genomes using NGS?
The key in NGS sequencing is the starting material. If the virus genome is isolated with high quality DNA/RNA, then NGS will provide high quality data. If the isolated DNA/RNA is degraded, then the sequencing result will also decrease in quality. Hope this answers your question!
thanks , very useful.
That's great to hear! :) Glad you found the video helpful.
OMG thank U so much!! IM really love that!! 🥰
You're welcome :) Glad you found the video helpful!
ciao ti seguo da quando la DNA polimerasi non era ancora stata scoperta , spero che leggiate il mio commento , vi mando un abbraccio 🦝😽😽
I woul really love to know how you make these powerful videos. I hope to start CZcams teaching in the future. I would really use a head-start from you with how to make these videos. I am a Biochemist so my videos will require some of these. Thank in advance
Hello Mene,
Sure thing! Feel free to use our content as your resources ;) Good luck with your youtube teaching channel!
Maybe you could have explained with all steps why the particular steps are taken. As somebody viewing this video with a limited understanding of NGS it's hard to follow what is going on at all and by the end of the video I still had no idea what this technique was about.
Hi BoBu,
Thank you for your comment. We will pass your feedback to our content production team.
Could you please tell me what are the impacts of NGS on the study of gene regulation?
czcams.com/video/XXDRRuQH-Rk/video.html&feature=share&fbclid=IwAR3i2m8gBtN40CqThutxi3jK1u86ZPhnaPXbauyEJvoss7tZDHrVgHVs0Rw
So, what is the difference between Sanger's sequencing and Sequencing by Synthesis?
Hi! We actually talk about the two in detail on our NGS Introduction Knowledge Base: old.abmgood.com/marketing/knowledge_base/next_generation_sequencing_introduction.php#SS (Look for the headings"Sanger Sequencing" and "Sequencing by Synthesis")
I want to invest in this
This video shall constitute a full essay for my final year exam in biotech lol
Haha, glad to know that our video is helpful! If you have any technical questions, we'll be happy to help answer them. We hope that you'll get an A+ in your class!
@@abmgood Jeez, I was here a year ago for my exam XD
can you please tell me
about the adapters???
+zunaira ehsan the adapters are short DNA sequences that need to be ligated to your library. Their function is to bind to their complementary sequence on the sequencer chip - you can learn more about them here: goo.gl/9u6SxW - let us know if you have any specific questions.
I have obtained 16S and ITS primers based metaseq data from a NGS company now how can I analyse that data??
Hi Aqleem,
There are many open source tools that are capable of analyzing 16S or ITS metagenomic data (E.g. CHIME and Mothur to name a few). Each comes with their own set of commands to perform the analysis you need to answer your question.
abm also offers bioinformatics analysis service for metagenomics data. Whether your sequencing data is from abm or from another sequencer, we would love to help with your data analytical needs. Please contact NGS@abmgood.com, and our technical staff will be happy to provide you with further details and assistance.
I have got Two files one for ITS files and the other for 16S files. It is metasequencing or metagenomic data. It is not based on simple gene identification, like species based identification. Therefore I cannot analyse the data,
Thanks for such a good video.
very good thank you so much
Thank you for watching! ;)
Can you define the Clusters what do they mean exactly?
Hi Rawand, thanks for watching and leaving your question! A cluster in this case, is a group of DNA that was amplified from a single original library fragment. Hope this helps!
I didn't find her to be too fast. Besides, if you need to, you can slow down the video a tad; there's no need to get bent out of shape. Very informational video! Thank you for your time and effort.
Hi Elise,
Thanks for watching and leaving a comment! We are glad you enjoyed our video - if you would like to learn more about NGS, you can check our our Knowledge Base: goo.gl/Ce0M4O
Hi Elise! Thank you for your kind comments! We're happy you enjoyed the video - let us know if you have any questions about the material presented.
The video is perfect because it explains things in a very synthetic way, I find the background music particularly relaxing so could you just give me the music that will save? It is perfect for studying.
Hi Daniel, thank you so much for the kind comment. Glad to hear that the video was helpful! Sorry, I'm not sure what is the song that we used here as the video was made several years ago. :(
@Daniel Hi, I know it's been 11 months but here it is. Capo productions - Daydreaming czcams.com/video/k2-LrO0Eupw/video.html
@@SofiannaMousti wow thank you very much
@@DANIEL_--_-_-_-_521 I'm happy to help! :)
What is mean by solid Sequencing method??
Hi Sudip, SOLiD sequencing is a type of Sequencing by Ligation method. You can read more about the SOLiD sequencing method on our knowledge base: goo.gl/o3gjDe
Is it covers BRACA 1 2 TESTS ALSO
Hello Arun,
BRCA1 and BRCA2 testing can be done via Next Generation Sequencing.
Good video, but it would be easier to understand if the woman talked slower.
Hi Hiske! Thanks for watching our video--we will certainly take your comment into consideration in our future videos. Please let us know if you have any questions about the material presented, we'd be glad to help!
video settings, speed
you can slow down or speed up the video the way you like.
@@ShytFerBraynes It doesn't help. The speaker eats words in some points. I am not native speaker and I can understand very well some sentences while other are just a confused sound. Even youtube can't generate proper subtitles!
@Albert Jackson and you are the kid who think to be smart. I guess it's matter of points of views... It would be nice and good for you as well if you try to stay in other people shoes.
Amazing video, thanks a lot. Pace could be a bit slower though.
Hey Argon, glad to hear that you enjoyed the video! We've been working on improving the pacing in our newer videos. If this helps, you can adjust the playback speed (under "Settings" on this video) to make it a bit slower.
thanks a lot
what is the primer in sanger squencing？ can you tell me
If you're looking to learn more about what primers are in general, the Wikipedia article on primers is actually a great starting point: en.wikipedia.org/wiki/Primer_(molecular_biology). The article talks about primers in Sanger Sequencing in this passage: "DNA sequencing is used to determine the nucleotides in a DNA strand. The Sanger chain termination method of sequencing uses a primer to start the chain reaction. It is worth noting that primers are not always for DNA synthesis, but can in fact be used by viral polymerases, e.g. influenza, for RNA synthesis." Let me know if this helps or if you have further questions!
Applied Biological Materials - abm In sanger squencing.the sequence determination using restriction fragments A12d and A14 as primers on the complementary strand of φX174 DNA. So, why to use them as primers. And in human genetic sequencing ,the sequences are all unknown. So we use sanger sequence to do it？how to finish the work
@@yafeiguo4149 Sorry, could you clarify your question? Is this question related to something specific to the project you are working on?
It is very fast paced !
Hi Peter,
Thanks for your comment! You can try slowing down the video's pace by adjusting your youtube viewing speed (check under the "Settings" or gear icon on the bottom right of the video).
Thank you very much.
This video was very helpful! But the correct name for the NGS system is "Illumina MiSeq" rather than "MySeq" at 2:21.
Hello Camila, thanks for pointing this out! We will make a note of this in the video description!
Thanks, it helped a lot.
+sphinxproduction That is excellent to hear! Thank you.
I have a confusion "What's different between 2nd Generation sequencing and next Generation sequencing ??"
Sanger and Maxam-Gilbert sequencing technologies (i.e. first generation) were the most common sequencing technologies until the development of massively parallel sequencing. To distinguish the new technologies from the old, they were called "next generation" or "second generation" sequencing (NGS) methods. Therefore when researchers talk about NGS, they are often referring to second generation sequencing. However, keep in mind that there might always be a "next" generation as new sequencing methods get developed. Therefore to be specific, it is best to refer to the different generations as "first", "second", or "third" instead of "next".
@@abmgood Thanks ❤️
Very helpful video, although it did make me want to play 2 Dots by the end
Hi Hannah, thanks for watching and glad you enjoyed our video! Let us know if you have any questions (or feel free to explore our knowledge base: goo.gl/Ce0M4O)
This is helpful video. Could you post the transcript here, please! Thank you so much ^^
Hi nhan, thanks for your comment - glad you enjoyed our video! You can actually find the transcript if you click on "More" (3 dots). There should be a "Transcript" option available.
??
I WANT A SUMMARY ABOUT THE PRINCIPLE OF NGS WITH THE REFERENCE PLEASE
-_-
Hi! Have you checked out our knowledge base article on the introduction to NGS? It is fully referenced. You can view it here: www.abmgood.com/marketing/knowledge_base/next_generation_sequencing_introduction.php
what is the maximum length (bp) of NGS your company can do?
The maximum read length we can sequence is 2x300bp, using MiSeq V3. Larger templates can undergo a tagmentation step first, i.e. transposases randomly cut the DNA template into short fragments ("tags") followed by addition of adaptors on either side of the cut points (ligation).
How on earth did they summarise NGS better than my biomedical professors?? They should have just made us watch this instead, geez. I now know why we have to use electrophoresis and chromatograms to interpret the results.
Also, the background music was relaxing and nice (like a commercial), ignore everyone else.
Hello,
We are very glad to hear that you found our video helpful! :) Stay tuned for upcoming videos as well! Also, please contact us at technical@abmgood.com if you have any questions. Cheers!
What about SMRT and nanopore?
Hi Sally, PacBio and Oxford nanopore are emerging sequencing technologies that are vastly different from Illumina sequencing and Sanger sequencing. We are planning on releasing a video describing these technologies as well. Please stay tuned!
A video would be great! I really enjoyed this one.
Thanks for your support! We currently have 4 more videos in our NGS playlist that go into more technical details as well as real-world applications based on case studies.
Sounds amazing! I look forward to them.
Mistake in the video, please correct -Sanger sequencing was developed in 1977, not 1900.
Hi Dilnora, thanks for pointing this out. The correction was previously added in as an annotation, but since CZcams has removed that feature, we went ahead and added the info in the video description.
jai swaminarayan
very helpful...but i quess, its no more possible to talk faster. with half the playback speed its very good.
Hello Juan,
We are sorry to hear that! I will definitely pass your feedback to our content production team. Thank you!
background music is really annoying! It seems that teasing me. Moreover, what is index1 reading and cluster indicate what? I think if you explain about the definition of terminology, people will be able to understand the process of NGS better!
Thanks for your comment. We'll definitely work on fixing the sound and music in our future videos so that they won't be so distracting! For your questions, could you clarify which part of the video you found confusing (or provide timestamps)?
Very informative video. Next time please ensure to reduce the volume of background score or music so that listeners can easily capture the speakers voice over the music.
Hi Aswathy, thanks for watching our video and glad you found it helpful! We are always looking to improve our audio quality - thanks for leaving your feedback!
❤
thanks for the good video but she talks too fast for me ! and the background music is disturbing.
Hi Narges, thanks for your comment! Yes, unfortunately, this was one of our earlier videos and we've been working on fixing those issues in our newer releases. If this helps, our videos have closed captions that you can turn on if you want to lower the volume, or you can also try slowing down the speed under the video's Settings > Speed option.
There are bad videos and then there is this video! Not sure if the viewer is supposed to listen to the speaker or the infernal music. Multiple viewers have made the comment on the music, but either the moderators are not reading the comments or they just don't care!
Hello Mitali,
We are really sorry to hear that! Indeed, we do care very much about our viewers feedback and have replied back to everyone's comment on the music. Unfortunately, we couldn't replace the old videos but you can always mute the video and turn on the caption instead if it bothers too much! :)
This video could be of interest for anyone looking for Next Generation Sequencing:
czcams.com/video/vkcTB2wKu3k/video.html
nauma ijaz ,I think you also watching the same video.....
The background music is a noise.
Please remove the background music, distracting and not needed.
Hi Desertfleur01, thanks for your comment! We've taken note for our future videos. If the music is too loud, we have close-captioning on this video which you can switch on! Thanks for watching!
@@abmgood I like the music lol
@@franciscos.2301 Glad to hear that :) Thanks for watching our video!
@@abmgood I like the music too!
@@samuelwallace4291 Thanks for watching, Samuel!
The background music is too loud and distracting
Hello there,
We are well aware of this issue but unfortunately we couldn't replace all the old videos.
However, we have made improvements on our latest videos so please check them out! :)
As an alternative solution, you can always mute the video and
turn the caption on if the background music bothers too much :)
Thank you!
Sanger Sequencing Developed in 1977
2:22
Illumina NGS is best