Illumina ChIP-Seq

aperera · 05-12-2011, 01:09 PM

What kits are people currently using to make Illumina multiplexed ChIP-Seq libraries? We are not able to get an answer from Illumina regarding when the TruSeq ChIP-Seq kits will be available or what suggestions they have in the meantime. We have tried using both TruSeq genomic kits and RNA-Seq kits with some luck. However, we are unable to get good libraries consistently. Any feedback/suggestions would be greatly apprecicated!

JHU-ChIPmaniac · 05-13-2011, 01:11 PM

I don't have an answer for you but I'm also interested in multiplexing ChIP-seq libraries. I was going to try the Tru-seq kit. What kind of problems are you running into?

Chipper · 05-13-2011, 03:06 PM

Bioo NEXTflex kits comes with indexed adapters.

RNAseqer · 05-17-2011, 06:44 PM

We're using their indexed adapters for ChIP-Seq, mRNA-Seq and Genomic sequencing. Heard from our sales rep that they are launching a 48 plex ChIP-Seq kit this month

Quote:

Originally Posted by Chipper

Bioo NEXTflex kits comes with indexed adapters.

aperera · 05-24-2011, 11:37 AM

We are seeing inconsistent quality of libraries when TruSeq kits are used. I am looking in to Bioo Scientific kits. Also heard that they will release ChIP-Seq kits next. Can not wait!!!

NextGenSeq · 05-25-2011, 01:42 PM

NuGen is releasing a low input Chip-Seq kit next month.

aperera · 06-13-2011, 10:20 AM

Has anyone tried the Millipore's Magna ChIP-Seq kit? We are planning on trying this soon and will share results when available.

ETHANol · 06-19-2011, 05:28 AM

Follow the Illumina ChiP-seq protocol, but buy the enzymes from NEB and the primers from IDT (or whoever you buy your oligos from) HPLC or PAGE purified.

See this thread for the primer sequences. http://seqanswers.com/forums/showthread.php?t=198

Just be careful one of the .pdf files that someone posts has a mistake in it. You can also get the primer sequences if you e-mail Illumina but just remember to phosphothiorylate as described in the Illumina thread.

For barcoding/multiplexing ChIP-seq, just add a barcode to the end of the primers as described in this article. And so as you would before.
http://www.ncbi.nlm.nih.gov/pubmed/19159457

Arabette · 06-22-2011, 12:18 PM

Quote:

Originally Posted by aperera

What kits are people currently using to make Illumina multiplexed ChIP-Seq libraries? We are not able to get an answer from Illumina regarding when the TruSeq ChIP-Seq kits will be available or what suggestions they have in the meantime. We have tried using both TruSeq genomic kits and RNA-Seq kits with some luck. However, we are unable to get good libraries consistently. Any feedback/suggestions would be greatly apprecicated!

At least you got some success to some extent! I'm about to try this tomorrow (I cannot wait more): I am going to use the TruSeq library prep kit for DNA and change the enzyme and reaction volumes to match the older, validated ChIP protocol and titrate the adapter to account for the much lower amont of material going into the library...is that what you did? Thanks in advance for your reply (possibly by tomorrow!)

aperera · 09-28-2011, 09:57 AM

Just a quick update...
We tried the Millipore Magna ChIP-Seq kit with the Bioo Scientific adapters, and obtained beautiful libraries. We have only constructed 10 libraries so far, and hope to generate more data points in the near future. Cost per library comes to about $50.

frigo · 12-14-2011, 08:02 AM

Hello,
I have to do a ChIP-Seq multiplexed library and I would like to have some feedback about using the Truseq library prep kit and the Bioo Scientific kits. Does someone tries these kits ?? Is there too much problems with the Truseq library prep kit?
About the Millipore's Magna ChIP-Seq kit, it's use to do a ChIP and a sequencing but I just have to do the sequencing from ChIP samples. Does someone knows other kits ?
Sorry for my bad english and thank you for your answers

ETHANol · 12-15-2011, 01:15 AM

http://seqanswers.com/forums/showthread.php?t=13093

I'd say go with the Bioo kit as it is essentially what I am doing. The TruSeq kit you will have to modify some, and if this is your first shot, perhaps you don't want to be working out your own modifications. But I would also consider using my protocol in the linked in the thread above as the Kapa polymerase is significantly more efficient then Phusion (side by side comparison in hands) and Phusion is significantly better then the TruSeq polymerase at least according to the advertising.

frigo · 12-15-2011, 02:19 AM

Thank you very much for your answer ETHANol !
yes this will be my first shot so I think I'll test the Bioo kit. In a second time I will maybe study your protocol if it can optimized the manipulation.
thank you

msheldon · 12-16-2011, 07:05 AM

We are using the Bioo ChIP Seq Kit and have gotten great sequencing results with both Chip DNA and regular genomic DNA. What is truly great is that we can finally multiplex our Chip samples...ahhhhh.

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05-12-2011, 01:09 PM	#1
aperera Member Location: Kansas City Join Date: Jan 2008 Posts: 23	Illumina ChIP-Seq What kits are people currently using to make Illumina multiplexed ChIP-Seq libraries? We are not able to get an answer from Illumina regarding when the TruSeq ChIP-Seq kits will be available or what suggestions they have in the meantime. We have tried using both TruSeq genomic kits and RNA-Seq kits with some luck. However, we are unable to get good libraries consistently. Any feedback/suggestions would be greatly apprecicated!

12-15-2011, 01:15 AM	#12
ETHANol Senior Member Location: Western Australia Join Date: Feb 2010 Posts: 308	http://seqanswers.com/forums/showthread.php?t=13093 I'd say go with the Bioo kit as it is essentially what I am doing. The TruSeq kit you will have to modify some, and if this is your first shot, perhaps you don't want to be working out your own modifications. But I would also consider using my protocol in the linked in the thread above as the Kapa polymerase is significantly more efficient then Phusion (side by side comparison in hands) and Phusion is significantly better then the TruSeq polymerase at least according to the advertising. __________________ -------------- Ethan

05-13-2011, 01:11 PM	#2
JHU-ChIPmaniac Member Location: Baltimore Join Date: Jul 2010 Posts: 21	I don't have an answer for you but I'm also interested in multiplexing ChIP-seq libraries. I was going to try the Tru-seq kit. What kind of problems are you running into?

05-13-2011, 03:06 PM	#3
Chipper Senior Member Location: Sweden Join Date: Mar 2008 Posts: 324	Bioo NEXTflex kits comes with indexed adapters.

05-24-2011, 11:37 AM	#5
aperera Member Location: Kansas City Join Date: Jan 2008 Posts: 23	We are seeing inconsistent quality of libraries when TruSeq kits are used. I am looking in to Bioo Scientific kits. Also heard that they will release ChIP-Seq kits next. Can not wait!!!

05-25-2011, 01:42 PM	#6
NextGenSeq Senior Member Location: USA Join Date: Apr 2009 Posts: 482	NuGen is releasing a low input Chip-Seq kit next month.

06-13-2011, 10:20 AM	#7
aperera Member Location: Kansas City Join Date: Jan 2008 Posts: 23	Has anyone tried the Millipore's Magna ChIP-Seq kit? We are planning on trying this soon and will share results when available.

06-19-2011, 05:28 AM	#8
ETHANol Senior Member Location: Western Australia Join Date: Feb 2010 Posts: 308	Follow the Illumina ChiP-seq protocol, but buy the enzymes from NEB and the primers from IDT (or whoever you buy your oligos from) HPLC or PAGE purified. See this thread for the primer sequences. http://seqanswers.com/forums/showthread.php?t=198 Just be careful one of the .pdf files that someone posts has a mistake in it. You can also get the primer sequences if you e-mail Illumina but just remember to phosphothiorylate as described in the Illumina thread. For barcoding/multiplexing ChIP-seq, just add a barcode to the end of the primers as described in this article. And so as you would before. http://www.ncbi.nlm.nih.gov/pubmed/19159457

09-28-2011, 09:57 AM	#10
aperera Member Location: Kansas City Join Date: Jan 2008 Posts: 23	Just a quick update... We tried the Millipore Magna ChIP-Seq kit with the Bioo Scientific adapters, and obtained beautiful libraries. We have only constructed 10 libraries so far, and hope to generate more data points in the near future. Cost per library comes to about $50.

12-14-2011, 08:02 AM	#11
frigo Junior Member Location: paris Join Date: Oct 2011 Posts: 6	Hello, I have to do a ChIP-Seq multiplexed library and I would like to have some feedback about using the Truseq library prep kit and the Bioo Scientific kits. Does someone tries these kits ?? Is there too much problems with the Truseq library prep kit? About the Millipore's Magna ChIP-Seq kit, it's use to do a ChIP and a sequencing but I just have to do the sequencing from ChIP samples. Does someone knows other kits ? Sorry for my bad english and thank you for your answers

12-15-2011, 02:19 AM	#13
frigo Junior Member Location: paris Join Date: Oct 2011 Posts: 6	Thank you very much for your answer ETHANol ! yes this will be my first shot so I think I'll test the Bioo kit. In a second time I will maybe study your protocol if it can optimized the manipulation. thank you

12-16-2011, 07:05 AM	#14
msheldon Member Location: NYU Join Date: Jul 2011 Posts: 15	We are using the Bioo ChIP Seq Kit and have gotten great sequencing results with both Chip DNA and regular genomic DNA. What is truly great is that we can finally multiplex our Chip samples...ahhhhh.