In this preliminary study, differentially expressed genes were investigated in cranial tissues from chickens with hereditary exencephaly using cDNA microarrays containing 1152 genes and expressed sequence tags (ESTs). Genes showing twofold or greater differences at P < 0.05 between affected and normal cranial cells were considered to be candidates for hereditary exencephaly in chicken. Eighteen ESTs (11 known genes/homologues) were upregulated and 108 ESTs (51 known genes/homologues) were downregulated. The EST AL584231 (ROS006C9), orthologous to human MTHFD1, a known candidate gene for human neural tube defects (NTDs), was expressed at the same level both in normal and affected chicken cranial tissues. ESTs AL584253 (ROS006F7, thioredoxin reductase 1) and AL585511 (ROS024H9, thioredoxin), both involved in NTD pathogenic pathways in mice, were downregulated and had mean ratios of 0.41 and 0.04 for expression in affected vs. normal cells respectively. Expression differences of these two ESTs were confirmed by quantitative real-time polymerase chain reaction. These data indicate that ESTs AL584253 and AL585511 are candidates for hereditary exencephaly in chickens.

Double-color fluorescence in situ hybridization was performed on chicken chromosomes using seven unique clones from the human chromosome 3-specific NotI linking libraries. Six of them (NL1-097, NL2-092, NL2-230, NLM- 007, NLM-118, and NLM-196) were located on the same chicken microchromosome and NL1-290 on another. Two chicken microchromosome GGA15-specific BAC clones, JE024F14 containing the IGVPS gene and JE020G17 contain- ing the ALDH1A1 gene, were cytogenetically mapped to the same microchromosome that carried the six NotI linking clones, allowing identification of this chromosome as GGA15. Two GGA14-specific clones, JE027C23 and JE014E08 con- taining the HBA gene cluster, were co-localized on the same microchromosome as NL1-290, suggesting that this chromo- some was GGA14. The results indicated that the human chro- mosomal region HSA3q13 → q23 is likely to be orthologous to GGA15 and GGA14. The breakpoint of evolutionary conser- vation of human and chicken chromosomes was detected on HSA3q13.3 → q23 between NL1-290, on the one hand, and six other NotI clones, on the other hand. Considering the available chicken-human comparative mapping data, another break- point appears to exist between the above NotI loci and four oth- er genes, TFRC, EIF4A2, SKIL and DHX36 located on HSA3q24 → qter and GGA9. Based on human sequences with- in the NotI clones, localization of the six new chicken coding sequences orthologous to the human/rodent genes was suggest- ed to be on GGA15 and one on GGA14. Microchromosomal location of seven NotI clones from the HSA3q21 T-band region can be considered as evidence in support of our hypothesis about the functional analogy of mammalian T-bands and avian microchromosomes.