Facilitated Large-Scale Sequence Validation Platform Using Tn5-Tagmented Cell Lysates

Byungjin Hwang, Sunghoon Heo, Namjin Cho, Hanna Seo, Duhee Bang

Research output: Contribution to journalArticle

Abstract

A typical molecular cloning procedure requires Sanger sequencing for sequence validation, which is cost-prohibitive and labor-intensive for large-scale clone analysis in genotype-phenotype studies. Here we present the cost-effective clone analysis platform TnClone, which uses next-generation sequencing based on Tn5 tagmentation to rapidly analyze a large number of clones from cell lysates. This method bypasses the extensive plasmid purification step. We also developed a user-friendly graphical user interface and provided general guidelines for conducting validation experiments. We tested our program with 1023 plasmids (222 from cell lysates and 801 from purified clones) and achieved 92% and 99.3% sensitivity with cell lysates and purified DNA, respectively. Our platform provides rapid turnaround with minimal hands-on time for secondary evaluation, as next-generation sequencing technology continues to evolve.

Original languageEnglish
Pages (from-to)596-600
Number of pages5
JournalACS Synthetic Biology
Volume8
Issue number3
DOIs
Publication statusPublished - 2019 Mar 15

Fingerprint

Plasmids
Clone Cells
Cloning
Graphical user interfaces
Purification
Costs
DNA
Personnel
Costs and Cost Analysis
Molecular Cloning
Experiments
Genotype
Guidelines
Technology
Phenotype

All Science Journal Classification (ASJC) codes

  • Biomedical Engineering
  • Biochemistry, Genetics and Molecular Biology (miscellaneous)

Cite this

Hwang, Byungjin ; Heo, Sunghoon ; Cho, Namjin ; Seo, Hanna ; Bang, Duhee. / Facilitated Large-Scale Sequence Validation Platform Using Tn5-Tagmented Cell Lysates. In: ACS Synthetic Biology. 2019 ; Vol. 8, No. 3. pp. 596-600.
@article{85223bf618e04236922ec9262973226d,
title = "Facilitated Large-Scale Sequence Validation Platform Using Tn5-Tagmented Cell Lysates",
abstract = "A typical molecular cloning procedure requires Sanger sequencing for sequence validation, which is cost-prohibitive and labor-intensive for large-scale clone analysis in genotype-phenotype studies. Here we present the cost-effective clone analysis platform TnClone, which uses next-generation sequencing based on Tn5 tagmentation to rapidly analyze a large number of clones from cell lysates. This method bypasses the extensive plasmid purification step. We also developed a user-friendly graphical user interface and provided general guidelines for conducting validation experiments. We tested our program with 1023 plasmids (222 from cell lysates and 801 from purified clones) and achieved 92{\%} and 99.3{\%} sensitivity with cell lysates and purified DNA, respectively. Our platform provides rapid turnaround with minimal hands-on time for secondary evaluation, as next-generation sequencing technology continues to evolve.",
author = "Byungjin Hwang and Sunghoon Heo and Namjin Cho and Hanna Seo and Duhee Bang",
year = "2019",
month = "3",
day = "15",
doi = "10.1021/acssynbio.8b00482",
language = "English",
volume = "8",
pages = "596--600",
journal = "ACS Synthetic Biology",
issn = "2161-5063",
publisher = "American Chemical Society",
number = "3",

}

Facilitated Large-Scale Sequence Validation Platform Using Tn5-Tagmented Cell Lysates. / Hwang, Byungjin; Heo, Sunghoon; Cho, Namjin; Seo, Hanna; Bang, Duhee.

In: ACS Synthetic Biology, Vol. 8, No. 3, 15.03.2019, p. 596-600.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Facilitated Large-Scale Sequence Validation Platform Using Tn5-Tagmented Cell Lysates

AU - Hwang, Byungjin

AU - Heo, Sunghoon

AU - Cho, Namjin

AU - Seo, Hanna

AU - Bang, Duhee

PY - 2019/3/15

Y1 - 2019/3/15

N2 - A typical molecular cloning procedure requires Sanger sequencing for sequence validation, which is cost-prohibitive and labor-intensive for large-scale clone analysis in genotype-phenotype studies. Here we present the cost-effective clone analysis platform TnClone, which uses next-generation sequencing based on Tn5 tagmentation to rapidly analyze a large number of clones from cell lysates. This method bypasses the extensive plasmid purification step. We also developed a user-friendly graphical user interface and provided general guidelines for conducting validation experiments. We tested our program with 1023 plasmids (222 from cell lysates and 801 from purified clones) and achieved 92% and 99.3% sensitivity with cell lysates and purified DNA, respectively. Our platform provides rapid turnaround with minimal hands-on time for secondary evaluation, as next-generation sequencing technology continues to evolve.

AB - A typical molecular cloning procedure requires Sanger sequencing for sequence validation, which is cost-prohibitive and labor-intensive for large-scale clone analysis in genotype-phenotype studies. Here we present the cost-effective clone analysis platform TnClone, which uses next-generation sequencing based on Tn5 tagmentation to rapidly analyze a large number of clones from cell lysates. This method bypasses the extensive plasmid purification step. We also developed a user-friendly graphical user interface and provided general guidelines for conducting validation experiments. We tested our program with 1023 plasmids (222 from cell lysates and 801 from purified clones) and achieved 92% and 99.3% sensitivity with cell lysates and purified DNA, respectively. Our platform provides rapid turnaround with minimal hands-on time for secondary evaluation, as next-generation sequencing technology continues to evolve.

UR - http://www.scopus.com/inward/record.url?scp=85062364359&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85062364359&partnerID=8YFLogxK

U2 - 10.1021/acssynbio.8b00482

DO - 10.1021/acssynbio.8b00482

M3 - Article

C2 - 30726053

AN - SCOPUS:85062364359

VL - 8

SP - 596

EP - 600

JO - ACS Synthetic Biology

JF - ACS Synthetic Biology

SN - 2161-5063

IS - 3

ER -