The cDNA for rhMPV/NL/1/00/E93K/S101P was constructed as previously described [17] . To generate rhMPV/ ΔM2-2 cDNA, two Swa I restriction sites were introduced into the SphI/ClaI subclone, which contained the SphI (nt 100) to Cla I (nt8678) fragment derived from rhMPV/NL/ 1/00/E93K/S101P, using a Quik change mutagenesis kit (Stratagene). The first SwaI site was positioned at nt5326, down stream of the M2-1 stop codon, and was generated with the primer 5' CAGTGAGCATGGTCCAATTTAAAT-TACTATAGAGG and its complement. The stop codon of M2-1 is in bold and the Swa I restriction site is underlined. The second SwaI site was positioned at nt5468, upstream of the native stop codon of M2-2, and was generated using the primer 5' CATAGAAATTATATATGTCAAGGCTTATT-TAAATTAG and its complement. Digestion with Swa I resulted in the removal of 142 nts of the M2-2 gene. The Stu I (nt4495) to Cla I (nt8693) fragment in the subclone, containing M2-1, SH, and G genes, was transferred into the full-length rhMPV/NL/1/00/E93K/S101P cDNA to form rhMPV/ΔM2-2 as depicted in Figure 1A . rhMPV/ ΔM2-2 cDNA plasmid used to recover virus was sequenced from nt4495 to nt8693 to confirm that no unexpected changes were generated by PCR during cloning.