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Survey of transcripts in the adult Drosophila brain.

Posey KL, Jones LB, Cerda R, Bajaj M, Huynh T, Hardin PE, Hardin SH - Genome Biol. (2001)

Bottom Line: From our initial characterization of 271 randomly chosen clones, we expect that approximately 11% of the clones in this library will identify transcribed sequences not found in expressed sequence tag databases.This work complements the Drosophila genome project by providing information that facilitates more complete annotation of the genomic sequence.This library should be a useful resource that will help in determining how basic brain functions operate at the molecular level.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology and Biochemistry, Institute of Molecular Biology, University of Houston, Houston, TX 77204-5513, USA.

ABSTRACT

Background: Classic methods of identifying genes involved in neural function include the laborious process of behavioral screening of mutagenized flies and then rescreening candidate lines for pleiotropic effects due to developmental defects. To accelerate the molecular analysis of brain function in Drosophila we constructed a cDNA library exclusively from adult brains. Our goal was to begin to develop a catalog of transcripts expressed in the brain. These transcripts are expected to contain a higher proportion of clones that are involved in neuronal function.

Results: The library contains approximately 6.75 million independent clones. From our initial characterization of 271 randomly chosen clones, we expect that approximately 11% of the clones in this library will identify transcribed sequences not found in expressed sequence tag databases. Furthermore, 15% of these 271 clones are not among the 13,601 predicted Drosophila genes.

Conclusions: Our analysis of this unique Drosophila brain library suggests that the number of genes may be underestimated in this organism. This work complements the Drosophila genome project by providing information that facilitates more complete annotation of the genomic sequence. This library should be a useful resource that will help in determining how basic brain functions operate at the molecular level.

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Scheme for classifying the Drosophila brain library clones.
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Figure 1: Scheme for classifying the Drosophila brain library clones.

Mentions: Desiccated brain tissue from adult Drosophila melanogaster was used to construct a library using the Stratagene Hybrid-Zap system. This library was designed for protein expression and, therefore, was constructed such that full-length cDNAs containing 5' untranslated regions are not likely to be present. The number of clones in the library and the size of the clones were used to assess the quality of the library. The number of clones in the primary library was determined by titering one of the five packaging reactions. The total number of clones in the primary library is 6.75 × 106 (that is, all five packaging reactions). From the analysis of the fully sequenced clones (141 novel and matched isolog clones reported in this study), the majority of the inserts (53%) were between 400 and 800 base pairs (511 base pairs ± 197 base pairs average deviation). Characterized clones from the library range between 139 to 1,746 base pairs (bp), including only 15 As of the poly(A) tail. The insert size for this library is as expected using the Stratagene Hybrid-Zap kit, given that the size-selection column retains DNA molecules larger than 200 bp) (Stratagene technical support, personal communication). Of the 283 clones that were either completely or end-tagged sequenced, approximately 4% (12 clones) did not contain an insert (Figure 1).


Survey of transcripts in the adult Drosophila brain.

Posey KL, Jones LB, Cerda R, Bajaj M, Huynh T, Hardin PE, Hardin SH - Genome Biol. (2001)

Scheme for classifying the Drosophila brain library clones.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC30707&req=5

Figure 1: Scheme for classifying the Drosophila brain library clones.
Mentions: Desiccated brain tissue from adult Drosophila melanogaster was used to construct a library using the Stratagene Hybrid-Zap system. This library was designed for protein expression and, therefore, was constructed such that full-length cDNAs containing 5' untranslated regions are not likely to be present. The number of clones in the library and the size of the clones were used to assess the quality of the library. The number of clones in the primary library was determined by titering one of the five packaging reactions. The total number of clones in the primary library is 6.75 × 106 (that is, all five packaging reactions). From the analysis of the fully sequenced clones (141 novel and matched isolog clones reported in this study), the majority of the inserts (53%) were between 400 and 800 base pairs (511 base pairs ± 197 base pairs average deviation). Characterized clones from the library range between 139 to 1,746 base pairs (bp), including only 15 As of the poly(A) tail. The insert size for this library is as expected using the Stratagene Hybrid-Zap kit, given that the size-selection column retains DNA molecules larger than 200 bp) (Stratagene technical support, personal communication). Of the 283 clones that were either completely or end-tagged sequenced, approximately 4% (12 clones) did not contain an insert (Figure 1).

Bottom Line: From our initial characterization of 271 randomly chosen clones, we expect that approximately 11% of the clones in this library will identify transcribed sequences not found in expressed sequence tag databases.This work complements the Drosophila genome project by providing information that facilitates more complete annotation of the genomic sequence.This library should be a useful resource that will help in determining how basic brain functions operate at the molecular level.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology and Biochemistry, Institute of Molecular Biology, University of Houston, Houston, TX 77204-5513, USA.

ABSTRACT

Background: Classic methods of identifying genes involved in neural function include the laborious process of behavioral screening of mutagenized flies and then rescreening candidate lines for pleiotropic effects due to developmental defects. To accelerate the molecular analysis of brain function in Drosophila we constructed a cDNA library exclusively from adult brains. Our goal was to begin to develop a catalog of transcripts expressed in the brain. These transcripts are expected to contain a higher proportion of clones that are involved in neuronal function.

Results: The library contains approximately 6.75 million independent clones. From our initial characterization of 271 randomly chosen clones, we expect that approximately 11% of the clones in this library will identify transcribed sequences not found in expressed sequence tag databases. Furthermore, 15% of these 271 clones are not among the 13,601 predicted Drosophila genes.

Conclusions: Our analysis of this unique Drosophila brain library suggests that the number of genes may be underestimated in this organism. This work complements the Drosophila genome project by providing information that facilitates more complete annotation of the genomic sequence. This library should be a useful resource that will help in determining how basic brain functions operate at the molecular level.

Show MeSH
Related in: MedlinePlus