The NIEHS Xenopus maternal EST project: interim analysis of the first 13,879 ESTs from unfertilized eggs

The sequencing of expressed sequence tags (ESTs) from Xenopus laevis has la gged behind efforts on many other common experimental organisms and man, pa rtly because of the pseudotetraploid nature of the Xenopus genome. Nonethel ess, large collections of Xenopus ESTs would be useful in gene discovery, o ligonucleotide-based knockout studies, gene chip analyses of normal and per turbed development, mapping studies in the related diploid frog X. tropical is, and for other reasons. We have created a normalized library of cDNAs fr om unfertilized Xenopus eggs. These cells contain all of the information ne cessary for the first several cell divisions in the early embryo, as well a s much of the information needed for embryonic pattern formation and cell f ate determination. To date, we have successfully sequenced 13,879 ESTs out of 16,607 attempts (83.6% success rate), with an average sequence read leng th of 508 bp. Using a fragment assembly program, these ESTs were assembled into 8,985 'contigs' comprised of up to 11 ESTs each. When these contigs we re used to search publicly available databases, 46.2% bore no relationship to protein or DNA sequences in the database at the significance level of 1e -6. Examination of a sample of 100 of the assembled contigs revealed that m ost (similar to 87%) were comprised of two apparent allelic variants. Expre ssion profiles of 16 of the most prominent contigs showed that 12 exhibited some degree of zygotic expression. These findings have implications for se quence-specific applications for Xenopus ESTs, particularly the use of alle le-specific oligonucleotides for knockout studies, differential hybridizati on techniques such as gene chip analysis, and the establishment of accurate nomenclature and databases for this species. (C) 2001 Published by Elsevie r Science B.V. All rights reserved.