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.
Thibault
Stephen T
ST
Exelixis, 170 Harbor Way, South San Francisco, California 94083-0511, USA. thibault@exelixis.com
Singer
Matthew A
MA
Miyazaki
Wesley Y
WY
Milash
Brett
B
Dompe
Nicholas A
NA
Singh
Carol M
CM
Buchholz
Ross
R
Demsky
Madelyn
M
Fawcett
Robert
R
Francis-Lang
Helen L
HL
Ryner
Lisa
L
Cheung
Lai Man
LM
Chong
Angela
A
Erickson
Cathy
C
Fisher
William W
WW
Greer
Kimberly
K
Hartouni
Stephanie R
SR
Howie
Elizabeth
E
Jakkula
Lakshmi
L
Joo
Daniel
D
Killpack
Keith
K
Laufer
Alex
A
Mazzotta
Julie
J
Smith
Ronald D
RD
Stevens
Lynn M
LM
Stuber
Christiana
C
Tan
Lory R
LR
Ventura
Richard
R
Woo
Alesa
A
Zakrajsek
Irena
I
Zhao
Lora
L
Chen
Feng
F
Swimmer
Candace
C
Kopczynski
Casey
C
Duyk
Geoffrey
G
Winberg
Margaret L
ML
Margolis
Jonathan
J
D000818:Animals; D004251:DNA Transposable Elements; D004331:Drosophila melanogaster / Q000235:genetics*; D017344:Genes, Insect; D016254:Mutagenesis, Insertional
D004251:DNA Transposable Elements