A Restriction Map of Cauliflower Mosaic Virus DNA (Strain PV 147). Mapping of the Cleavage Sites of HhaI, SacI, AvaI, PvuII, PstI, XbaI, EcoRI, BglII, HincII, HpaII and HindII + III

Abstract

The virion-extracted DNA ( M , 5 x lo6) of cauliflower mosaic virus (CaMV) has three singlestranded interruptions. The mapping of this DNA using eleven restriction endonucleases (HhaI, S a d , AvaI, PvuII, PstI, XbaI, EcoRI, BglII, HincII, HpaII and HindII + 111) is reported here. The existence of the three single-stranded breaks complicates the identification and the molecular weight determination of fragments produced by HpaII, HindIII and HindII + 111. Indeed the electrophoretic mobility of some fragments in which a single-stranded discontinuity is located is modified, and the fluorescence of ethidium bromide complexed with these fragments is reduced as compared to that observed for the other fragments existing in a molar ratio. These drawbacks were overcome by performing experiments of nick-translation of CaMV DNA with Eschevichia coli DNA polymerase I. From the data it follows that the CaMV DNA molecule bears 1 site for HhaI and Sac& 2 for AvaI and PvuII, 3 for PstI, 4 for XbaI, 5 for EcoRI, 6 for BglII and HincII, 11 for Abbreviations. CaMV, cauliflower mosaic virus; SV40, simian virus 40.

The complete nucleotide sequence of an infectious clone of cauliflower mosaic virus by M13mp7 shotgun sequencing

“…We elected to sequence an infectious clone rather than a preparation of viral DNA for two reasons. First, restriction endonuclease cleavage patterns of viral DNA extracted from infected plants exhibit a range of minor fragments, some of which may be attributable to variability in sequence among the population of molecules (15)(16)(17). Secondly, CaMV viral DNA contains three single strand interruptions with an unusual structure (18,14) which may cause difficulties during cloning (19).…”