BioMed Research International

Methodology Report

Use of Recombination-Mediated Genetic Engineering for Construction of Rescue Human Cytomegalovirus Bacterial Artificial Chromosome Clones

Table 1

Table of primers used for PCR. ToledoUL132KanF and ToledoUL151KanR: primers to amplify kan^R cassette for insertion of positive antibiotic selection marker and simultaneous deletion mutant construction. ZeoInsertF and ZeoInsertR: amplification of zeo^R cassette for insertion of positive antibiotic selection marker insertion within region IV between UL 130 and UL 149. pUC19-IVCap_F and pUC19-IVCap_R: amplification of plasmid cassette for region IV capture for rescue clone construction.


Primer	Sequence (5′ 3′)	Function of primers

ToledoUL132KanF	CGCGGACATAGCAAGAAATCCA	Amplify kan^Rcassette for antibiotic selection and region IV deletion mutant construction
	CGTCGCCACATCTCGAGAGCTCT
	TGTTGGCTAGTGCGTA
ToledoUL151KanR	CGACCAGCGCTTTGTGCGCGCT
	GCCTGTGCGTGTCGTCCCTCTGC
	CAGTGTTACAACCAA

ZeoInsertF	GTCCGGCAGGATAGCGGTTAAG	Amplify zeo^R cassette for antibiotic selection within region IV between UL 130 and UL 149
	GATTCGGTGCTAAGGCCGCATG
	GCCGCGGATGGATCC
ZeoInsertR	TATCTGCGTGGGTCTAATCATGG
	GTGTCACCGTGATCGCGGCCGC
	ACTAGTGATAGATCT

pUC19-IVCap_F	CATTCAGGCGCGCCGGTAGTGT	Amplify plasmid cassette for region IV capture into puc19 plasmid for rescue clone generation
	GTACAAAGGGAGGCGTGCTCAC
	GGCCCGCAACCCGGGTACCGAG
	CTCGAAT
pUC19-IVCap_R	TACTCAGGCCGGCCATCAAAAC
	GCGAGCCCATATCGCCGCCATC
	ATTGTAATCAGATGTGTGAAATT
	GTTATCC