Hello everyone, has anyone tried to modify the amount of tranposome during tagmantation using Nextera? We have already modified the protocol with good results concerning tagmentation time, but we are now considering of decreasing TD and I am worried if this would work....Does anyone know if the Rd1 and Rd2 seq primers are in the TD tube or in the TDE tube (which we do not plan to decrease)?
Seqanswers Leaderboard Ad
Collapse
Announcement
Collapse
No announcement yet.
X
-
I do not know what your aim is to modify the Illumina protocol. TD is 2x tagmentation buffer and if you are planning to modify its amount you will need to proportionally modify your reaction volume. TDE tube contains transposon complex which inserts 19 bp transposase recognition sequences appended with 14-15 nt overhangs to DNA in tagmentation reaction that are used during PCR to add sequencing and flow cell binding motives and index sequences. R1 and R2 sequencing primers come with sequencing kit (MiSeq) or as separate kit which are added to SBS reagents in HiSeq systems.
-
Decreasing TDE without any other modification will results in larger fragments. If you reduce TDE only , then you will not have enough buffer to use it in other reactions. So, if you are aiming for larger fragments it would make more sense to reduce reaction volume (TD, TDE and input DNA) and keep the input DNA amount the same.
Comment
Latest Articles
Collapse
-
by seqadmin
The field of epigenetics has traditionally concentrated more on DNA and how changes like methylation and phosphorylation of histones impact gene expression and regulation. However, our increased understanding of RNA modifications and their importance in cellular processes has led to a rise in epitranscriptomics research. “Epitranscriptomics brings together the concepts of epigenetics and gene expression,” explained Adrien Leger, PhD, Principal Research Scientist...-
Channel: Articles
04-22-2024, 07:01 AM -
-
by seqadmin
Proteins are often described as the workhorses of the cell, and identifying their sequences is key to understanding their role in biological processes and disease. Currently, the most common technique used to determine protein sequences is mass spectrometry. While still a valuable tool, mass spectrometry faces several limitations and requires a highly experienced scientist familiar with the equipment to operate it. Additionally, other proteomic methods, like affinity assays, are constrained...-
Channel: Articles
04-04-2024, 04:25 PM -
ad_right_rmr
Collapse
News
Collapse
Topics | Statistics | Last Post | ||
---|---|---|---|---|
Started by seqadmin, 04-25-2024, 11:49 AM
|
0 responses
18 views
0 likes
|
Last Post
by seqadmin
04-25-2024, 11:49 AM
|
||
Started by seqadmin, 04-24-2024, 08:47 AM
|
0 responses
17 views
0 likes
|
Last Post
by seqadmin
04-24-2024, 08:47 AM
|
||
Started by seqadmin, 04-11-2024, 12:08 PM
|
0 responses
62 views
0 likes
|
Last Post
by seqadmin
04-11-2024, 12:08 PM
|
||
Started by seqadmin, 04-10-2024, 10:19 PM
|
0 responses
60 views
0 likes
|
Last Post
by seqadmin
04-10-2024, 10:19 PM
|
Comment