The complete nucleotide sequence of the hemagglutinin (HA) gene of a type B influenza virus (B/Lee/40) was obtained by using cloned cDNA derived from the RNA segment. The gene is 1,882 nucleotides long and can code for a protein precursor of 584 amino acids. Structural features common to type A virus HAs are also conserved in the B virus HA. These include a hydrophobic signal peptide, hydrophobic NH2 and COOH termini of the HA2 subunit, and a HA1/HA2 cleavage site involving an arginine residue. The sequence of the B HA gene and its deduced amino acid sequence were compared to those ofa type A influenza virus (A/PR/8/34). When these two genes were aligned, it was found that 24% of the amino acids in the HAI subunits and 39% of the amino acids in the HA2 subunits are conserved. This degree of relatedness between type B virus and type A virus HAs (intertypic comparison) is similar to the homologies observed among certain type A virus HAs (intratypic comparison). A close evolutionary relationship is therefore suggested between the HAs of type A and type B influenza viruses.

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A portion of Moloney murine sarcoma virus DNA which is repeated at both ends of the provirus has been sequences. The nucleotide sequence, together with hybridization data obtained with in vitro pulse-labelled nascent viral RNA, indicate that initiation and termination of RNA synthesis occur within that region of the proviral DNA. A model for transcriptional readthrough of termination signals during RNA synthesis in this system is suggested.

Moloney murine sarcoma virions synthesize discrete DNA products in vitro which closely resemble those found in vivo shortly after infection. These in vitro products have been isolated by electrophoresis and mapped with restriction endonucleases. In addition to the full-genome-length 6-kilobase pair linear DNA, a 5.4-kilobase pair circular DNA molecule, an incomplete linear DNA molecule, and a 600-base pair molecule were detected. The 6-kilobase pair DNA contained a 600-base pair direct terminal repeat which was missing from the circular form and was partially represented on the incomplete linear DNA molecule. The 600base pair DNA contained sequences which were present in the 600-base pair direct repeat on the 6-kilobase pair DNA. The order of synthesis and the structure of these molecules detected in the in vitro reaction suggest that they are crucial intermediates in the formation ofthe final product ofin vitro reverse transcription. A model which accounts for the synthesis of all of these molecules during the initial stages of viral replication is suggested.

Moloney murine sarcoma virus (MSV) virions incubated under optimal conditions were shown to support extensive synthesis of double-stranded DNA. The major product, a 5950-base-pair (6-kilobase-pair DNA) double-stranded DNA, was characterized by cleavage with restriction endonucleases and shown to contain a 600-nucleotide-ong direct repeat at both ends of the MSV genome. Linear DNA molecules made in vivo shortly after infection were compared to the linear doublestranded DNA synthesized in vitro. The restriction maps of both viral DNA products were indistinguishable. The 600-base-pair repeat results in a progeny DNA molecule that is longer than the parental MSV genomic RNA. The generation of this repeat must involve a mechanism that allows the viral reverse transcriptase (RNA-dependent DNA nucleotidyltransferase) to copy 5'-and 3'-terminal genomic (+) Synthesis of DNA in Detergent-Disrupted Virions. Virions at a concentration of 5 mg/ml were disrupted with Triton X-100 (final concentration 0.028-0.030%) in a reaction mixture containing 50 mM Tris-HCI (pH 7.5), 20 mM dithiothreitol, 2mM MgCl2, and the four dNTPs at 1 mM each. DNA was labeled with [a-32P]dNTPs (Amersham or New England Nuclear) at a specific activity of 5-50 cpm/pmol or with 125I-labeled dCTP, made by a modification of the method of Commerford (5, 10). Incubations were at 37"C for 16-18 hr. These conditions optimized the synthesis of full-length double-stranded products.The DNA products were extracted with phenol saturated wth 50 mM Tris-HCI, pH 7.5/0.1 M NaCl/1 mM EDTA/0.1% sodium dodecyl sulfate (NaDodSO4) and precipitated twice with 2 vol of ethanol. This material was digested with an excess of nuclease S1 (Sigma, 5 units per ,ug of DNA) in 100 mM sodium acetate buffer, pH 4.5/0.2 M NaCI/1 mM ZnCl2 at 37°C for 30 min.Isolation and Purification of Viral DNA. Full-length viral dsDNA was isolated by electrophoresis on 1.5% neutral agarose (Sea Kem, HGT) horizontal slab gels in 50 mM borate/i mM EDTA at pH 8 at 150 V for about 16 hr. Restriction fragments of T7 DNA (11) or phage a3 replicative form II [32PJDNA served as molecular weight markers. The DNA was stained with ethidium bromide (0.1 ,ug/ml in 0.25 M ammonium acetate) for 30 min and visualized by UV illumination, and the bands were cut out. The DNA was eluted by crushing the agarose in 0.5 M ammonium acetate/10 mM EDTA/0.1% NaDodSO4, incubating for 16 hr at 370C, and separating the agarose from the DNA by centrifuging the mixture over silanized glass wool in a micropipet tip. The DNA was precipitated twice with 2 vol of ethanol at -70°C and collected by centrifugation. When necessary, DNA preparations were further purified by Bio-Gel A-5M chromatography in 4 ml of Quik-Sep (Isolab, Akron, OH) columns with 50 mM Tris-HCI/0. 1 M NaCI/2.5 mM EDTA/ 0.1% NaDodSO4/10% glycerol/0.01% diethylpyrocarbonate as solvent, precipitated twice with ethanol, and stored in 10 mM The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby...