To resolve the genome structure of these retroviral elements, we have determined the complete sequence of two proviral clones of EAV-HP from a line N chicken genomic DNA yeast artificial chromosome library and from a meat-type chicken line 21 lambda library. The EAV-HP sequences from the two lines were 98% identical and had a typical provirus structure. The two EAV-HP clones showed identical large deletions spanning part of the gag, the entire pol, and part of the env genes. The env region of the EAV-HP clones was 97% identical to the env sequence of HPRS-103, the prototype subgroup J ALV. The 5 region of EAV-HP comprising the R and U5 regions of the long terminal repeat (LTR), the untranslated leader, and the 5 end of the putative gag region were 97% identical to the avian retrotransposon sequence, ART-CH. The remaining gag sequence shared less than 60% identity with other ALV sequences. The U3 region of the LTR was distinct from those of other retroviruses but contained some of the conserved motifs required for functioning as a promoter. To examine the ability of this endogenous retroviral LTR to function as a transcriptional promoter, the EAV-HP and HPRS-103 LTR U3 regions were compared in a luciferase reporter gene assay. The low luciferase activity detected with the EAV-HP LTR U3 constructs, at levels close to those observed for a control vector lacking the promoter or enhancer elements, suggested that these elements function as a weak promoter, possibly accounting for their low expression levels in chicken embryos.Endogenous retrovirus (ERV) sequences inherited as Mendelian genes have been recognized in most of the vertebrate genomes. In the chicken genome, four different families of ERVs have been identified. The CR1 (chicken repeat 1) element, a short interspersed repetitive DNA element belonging to the non-long terminal repeat (LTR) class of retrotransposons, forms one of these families (15). The majority of these elements, existing as approximately 7,000 to 20,000 repeats per haploid genome, have a common 3Ј end but show variable 5Ј truncations, with a few elements containing open reading frames encoding reverse transcriptase (13). CR1 elements have been identified in several avian and reptilian species, demonstrating that they are ancient sequences that arose before the divergence of birds and reptiles (40).The second family of chicken retrovirus-like elements, with the best-characterized ev loci, are the avian leukosis virus (ALV) subgroup E (ALV-E) ERVs. There are over 22 ev loci documented in layer-type birds and probably more in meattype breeds (17), with an average of 5 loci in each bird (33). Although the majority of the ev loci are defective, some of them encode infectious ERVs closely related to the ALVs (reviewed in reference 18). The ev2 locus, for example, encodes Rous-associated virus-0, the prototype of ALV-E (27). The ev loci are considered to represent recent germ line integrations because of their (i) low copy numbers, (ii) segregation in the population, and (iii) distribution restrict...

A rapid method is described for non isotopic in situ mapping of single copy genes directly on G-banded chromosomes by "one-step" regular light microscopy. It is based on hybridizing biotinylated probes to metaphase chromosomes. Biotin residues are detected by rabbit antibiotin antibody and anti-rabbit Ig labelled with peroxidase or colloidal gold. The peroxidase reaction product or colloidal gold signals are amplified by silver precipitation. The final product is a black silver dot at the gene locus on a purple G-banded chromosome. N-ras and alpha-1-antitrypsin genes have been mapped using plasmids with inserts of 1.5 and 1.3kb to 1p13.1 and the junction of 14q31/32 respectively. The signal to noise ratio in these experiments ranged from 32:1-46:1. This technology is at least as sensitive as radioisotopic in situ hybridization and gives results within 1 day of hybridization and has much better resolution. Additionally, genes are visualized by regular light microscopy without specialized techniques such as reflection contrast, fluorescence or phase microscopy. This methodology should facilitate more precise chromosomal gene localization.

Avian leukosis virus of subgroup J (ALV-J), isolated in the late 1980s, predominantly causes myelocytic myeloid leukosis in meat-type chickens. In the past few years, we have observed the occurrence of lesions indicative of erythroblastosis in ALV-J-infected flocks and, in this paper, we report the isolation of ALV-J strains from such flocks. Three of these isolates were acutely transforming viruses, as shown by their ability to transform bone marrow cell cultures. The bone marrow cultures transformed by these virus isolates were very similar to the myeloid cells transformed by the ALV-J strain 966. However, the infection of meat-type chickens with these isolates either as embryos or as 1-day-old chicks resulted in the induction of erythroblastosis as well as myelocytomatosis. Other histopathological changes observed in the inoculated birds included neoplastic lesions such as cholangioma and testicular cell tumour, and non-neoplastic lesions such as lymphomyeloid hyperplasia. This report demonstrates that highly oncogenic ALV-J, capable of inducing a different spectrum of disease other than the widely reported myelocytomatosis, could be established in naturally infected flocks.

Summary The origin and cell lineage of stromal cells in the bone marrow is uncertain. Whether a common stem cell exists for both haemopoietic and stromal cells or whether these cell lines arise from distinct stem cells is unknown. Using in situ hybridisation for detection of the Y chromosome, we have examined histological sections of bone marrow from seven patients who received marrow transplants from HLA-matched donors of the opposite sex. Stromal cells (adipocytes, fibroblasts, endothelial cells, osteoblasts and osteocytes) were identified in these recipients as being of host origin. This result is consistent with the concept of a distinct origin and separate cell lineage for cells of the haemopoietic and stromal systems. It also shows that engraftment of marrow stromal cell precursors does not occur and that host stromal cells survive conditioning regimens for marrow transplantation. With the exception of one case, with a markedly hypocellular marrow, mixed chimaerism was seen in haemopoietic cells, indicating that this is not a rare event after marrow transplantation.