A complementary transposon tool kit for Drosophila melanogaster using P and piggyBac.
With the availability of complete genome sequence for Drosophila melanogaster, one of the next strategic goals for fly researchers is a complete gene knockout collection. The P-element transposon, the workhorse of D. melanogaster molecular genetics, has a pronounced nonrandom insertion spectrum. It has been estimated that 87% saturation of the approximately 13,500-gene complement of D. melanogaster might require generating and analyzing up to 150,000 insertions. We describe specific improvements to the lepidopteran transposon piggyBac and the P element that enabled us to tag and disrupt genes in D. melanogaster more efficiently. We generated over 29,000 inserts resulting in 53% gene saturation and a more diverse collection of phenotypically stronger insertional alleles. We found that piggyBac has distinct global and local gene-tagging behavior from that of P elements. Notably, piggyBac excisions from the germ line are nearly always precise, piggyBac does not share chromosomal hotspots associated with P and piggyBac is more effective at gene disruption because it lacks the P bias for insertion in 5' regulatory sequences.
Exelixis, 170 Harbor Way, South San Francisco, California 94083-0511, USA. thibault@exelixis.com
Stephen T
ST
Thibault
Matthew A
MA
Singer
Wesley Y
WY
Miyazaki
Brett
B
Milash
Nicholas A
NA
Dompe
Carol M
CM
Singh
Ross
R
Buchholz
Madelyn
M
Demsky
Robert
R
Fawcett
Helen L
HL
Francis-Lang
Lisa
L
Ryner
Lai Man
LM
Cheung
Angela
A
Chong
Cathy
C
Erickson
William W
WW
Fisher
Kimberly
K
Greer
Stephanie R
SR
Hartouni
Elizabeth
E
Howie
Lakshmi
L
Jakkula
Daniel
D
Joo
Keith
K
Killpack
Alex
A
Laufer
Julie
J
Mazzotta
Ronald D
RD
Smith
Lynn M
LM
Stevens
Christiana
C
Stuber
Lory R
LR
Tan
Richard
R
Ventura
Alesa
A
Woo
Irena
I
Zakrajsek
Lora
L
Zhao
Feng
F
Chen
Candace
C
Swimmer
Casey
C
Kopczynski
Geoffrey
G
Duyk
Margaret L
ML
Winberg
Jonathan
J
Margolis
D000818:Animals; D004251:DNA Transposable Elements; D004331:Drosophila melanogaster / Q000235:genetics*; D017344:Genes, Insect; D016254:Mutagenesis, Insertional
D004251:DNA Transposable Elements