Header Leaderboard Ad

Collapse

Assessing evenness of pooled plasmid library

Collapse

Announcement

Collapse
No announcement yet.
X
 
  • Filter
  • Time
  • Show
Clear All
new posts

  • Assessing evenness of pooled plasmid library

    Hello,

    I am wondering what is the best way to assess the evenness of a pooled plasmid library.

    I have both the original library consisting of an equimolar pool of individual DNA constructs, and I have a maxi DNA prep of the pooled library that was propagated in E. coli (i.e. E. coli was transformed with the original library).

    I have sequenced both samples on the NextSeq. I would like to asses both library evenness and any change that occurred as a result of propagating the library in E. coli.

    I know that this kind of analysis is often done for CRISPR GeCKO libraries (gRNA libraries), but my library is different because it is a gene library where the genes have a range of lengths. So I presume that the read counts will need to be normalised to the gene lengths.

    I was thinking of using a normalised read count like TPM or similar, since I understand that measure accounts for gene length.

    And then, once I have the normalised read count for each construct in the library, I wonder what would be the best algorithm to check for differences between the two samples (i.e. change due to propagation in E. coli).

    Any advice would be greatly appreciated.

    Thank you.
    Last edited by splot; 12-10-2021, 03:10 AM.

Latest Articles

Collapse

  • seqadmin
    Improved Targeted Sequencing: A Comprehensive Guide to Amplicon Sequencing
    by seqadmin



    Amplicon sequencing is a targeted approach that allows researchers to investigate specific regions of the genome. This technique is routinely used in applications such as variant identification, clinical research, and infectious disease surveillance. The amplicon sequencing process begins by designing primers that flank the regions of interest. The DNA sequences are then amplified through PCR (typically multiplex PCR) to produce amplicons complementary to the targets. RNA targets...
    03-21-2023, 01:49 PM
  • seqadmin
    Targeted Sequencing: Choosing Between Hybridization Capture and Amplicon Sequencing
    by seqadmin




    Targeted sequencing is an effective way to sequence and analyze specific genomic regions of interest. This method enables researchers to focus their efforts on their desired targets, as opposed to other methods like whole genome sequencing that involve the sequencing of total DNA. Utilizing targeted sequencing is an attractive option for many researchers because it is often faster, more cost-effective, and only generates applicable data. While there are many approaches...
    03-10-2023, 05:31 AM

ad_right_rmr

Collapse

News

Collapse

Topics Statistics Last Post
Started by seqadmin, 03-24-2023, 02:45 PM
0 responses
14 views
0 likes
Last Post seqadmin  
Started by seqadmin, 03-22-2023, 12:26 PM
0 responses
15 views
0 likes
Last Post seqadmin  
Started by seqadmin, 03-17-2023, 12:32 PM
0 responses
17 views
0 likes
Last Post seqadmin  
Started by seqadmin, 03-15-2023, 12:42 PM
0 responses
23 views
0 likes
Last Post seqadmin  
Working...
X