pELMO, an optimised in-house cloning vector

Andrea E. Ramos, Claudia Marina Muñoz, Darwin A. Moreno-Pérez, Manuel A. Patarroyo

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

DNA cloning is an essential tool regarding DNA recombinant technology as it allows the replication of foreign DNA fragments within a cell. pELMO was here constructed as an in-house cloning vector for rapid and low-cost PCR product propagation; it is an optimally designed vector containing the ccdB killer gene from the pDONR 221 plasmid, cloned into the pUC18 vector’s multiple cloning site (Thermo Scientific). The ccdB killer gene has a cleavage site (CCC/GGG) for the SmaI restriction enzyme which is used for vector linearisation and cloning blunt-ended products. pELMO transformation efficiency was evaluated with different sized inserts and its cloning efficiency was compared to that of the pGEM-T Easy commercial vector. The highest pELMO transformation efficiency was observed for ~500 bp DNA fragments; pELMO vector had higher cloning efficiency for all insert sizes tested. In-house and commercial vector cloned insert reads after sequencing were similar thus highlighting that sequencing primers were designed and localised appropriately. pELMO is thus proposed as a practical alternative for in-house cloning of PCR products in molecular biology laboratories.

Original languageEnglish (US)
Article number26
JournalAMB Express
Volume7
Issue number1
DOIs
StatePublished - Dec 1 2017

Fingerprint

Genetic Vectors
Organism Cloning
Polymerase Chain Reaction
Recombinant DNA
DNA
DNA Replication
Genes
Molecular Biology
Plasmids
Technology
Costs and Cost Analysis
Enzymes

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Applied Microbiology and Biotechnology

Cite this

Ramos, Andrea E. ; Muñoz, Claudia Marina ; Moreno-Pérez, Darwin A. ; Patarroyo, Manuel A. / pELMO, an optimised in-house cloning vector. In: AMB Express. 2017 ; Vol. 7, No. 1.
@article{6808dfe6fc7442ab91afb4dd1a3b7f79,
title = "pELMO, an optimised in-house cloning vector",
abstract = "DNA cloning is an essential tool regarding DNA recombinant technology as it allows the replication of foreign DNA fragments within a cell. pELMO was here constructed as an in-house cloning vector for rapid and low-cost PCR product propagation; it is an optimally designed vector containing the ccdB killer gene from the pDONR 221 plasmid, cloned into the pUC18 vector’s multiple cloning site (Thermo Scientific). The ccdB killer gene has a cleavage site (CCC/GGG) for the SmaI restriction enzyme which is used for vector linearisation and cloning blunt-ended products. pELMO transformation efficiency was evaluated with different sized inserts and its cloning efficiency was compared to that of the pGEM-T Easy commercial vector. The highest pELMO transformation efficiency was observed for ~500 bp DNA fragments; pELMO vector had higher cloning efficiency for all insert sizes tested. In-house and commercial vector cloned insert reads after sequencing were similar thus highlighting that sequencing primers were designed and localised appropriately. pELMO is thus proposed as a practical alternative for in-house cloning of PCR products in molecular biology laboratories.",
author = "Ramos, {Andrea E.} and Mu{\~n}oz, {Claudia Marina} and Moreno-P{\'e}rez, {Darwin A.} and Patarroyo, {Manuel A.}",
year = "2017",
month = "12",
day = "1",
doi = "10.1186/s13568-017-0324-2",
language = "English (US)",
volume = "7",
journal = "AMB Express",
issn = "2191-0855",
publisher = "Springer",
number = "1",

}

pELMO, an optimised in-house cloning vector. / Ramos, Andrea E.; Muñoz, Claudia Marina; Moreno-Pérez, Darwin A.; Patarroyo, Manuel A.

In: AMB Express, Vol. 7, No. 1, 26, 01.12.2017.

Research output: Contribution to journalArticle

TY - JOUR

T1 - pELMO, an optimised in-house cloning vector

AU - Ramos, Andrea E.

AU - Muñoz, Claudia Marina

AU - Moreno-Pérez, Darwin A.

AU - Patarroyo, Manuel A.

PY - 2017/12/1

Y1 - 2017/12/1

N2 - DNA cloning is an essential tool regarding DNA recombinant technology as it allows the replication of foreign DNA fragments within a cell. pELMO was here constructed as an in-house cloning vector for rapid and low-cost PCR product propagation; it is an optimally designed vector containing the ccdB killer gene from the pDONR 221 plasmid, cloned into the pUC18 vector’s multiple cloning site (Thermo Scientific). The ccdB killer gene has a cleavage site (CCC/GGG) for the SmaI restriction enzyme which is used for vector linearisation and cloning blunt-ended products. pELMO transformation efficiency was evaluated with different sized inserts and its cloning efficiency was compared to that of the pGEM-T Easy commercial vector. The highest pELMO transformation efficiency was observed for ~500 bp DNA fragments; pELMO vector had higher cloning efficiency for all insert sizes tested. In-house and commercial vector cloned insert reads after sequencing were similar thus highlighting that sequencing primers were designed and localised appropriately. pELMO is thus proposed as a practical alternative for in-house cloning of PCR products in molecular biology laboratories.

AB - DNA cloning is an essential tool regarding DNA recombinant technology as it allows the replication of foreign DNA fragments within a cell. pELMO was here constructed as an in-house cloning vector for rapid and low-cost PCR product propagation; it is an optimally designed vector containing the ccdB killer gene from the pDONR 221 plasmid, cloned into the pUC18 vector’s multiple cloning site (Thermo Scientific). The ccdB killer gene has a cleavage site (CCC/GGG) for the SmaI restriction enzyme which is used for vector linearisation and cloning blunt-ended products. pELMO transformation efficiency was evaluated with different sized inserts and its cloning efficiency was compared to that of the pGEM-T Easy commercial vector. The highest pELMO transformation efficiency was observed for ~500 bp DNA fragments; pELMO vector had higher cloning efficiency for all insert sizes tested. In-house and commercial vector cloned insert reads after sequencing were similar thus highlighting that sequencing primers were designed and localised appropriately. pELMO is thus proposed as a practical alternative for in-house cloning of PCR products in molecular biology laboratories.

UR - http://www.scopus.com/inward/record.url?scp=85010898426&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85010898426&partnerID=8YFLogxK

U2 - 10.1186/s13568-017-0324-2

DO - 10.1186/s13568-017-0324-2

M3 - Article

C2 - 28116699

AN - SCOPUS:85010898426

VL - 7

JO - AMB Express

JF - AMB Express

SN - 2191-0855

IS - 1

M1 - 26

ER -