PhiC31 integrase-mediated transgenesis systems
PhiC31 integrase-mediated transgenesis systems are based on the site-specific bacteriophage PhiC31 integrase which mediates sequence-directed, irreversible and highly efficient integration between a bacterial attachment site (attB) and a phage attachment site (attP). Injecting plasmid containing attB site and white marker into attP-containing docking site strain(s) with PhiC31 activity makes the resultant stable w+ transformants containing your gene-of-interest between attL and attR sites (irreversible).
We use M{vas-int.Dm}ZH-2A as the genomic PhiC31 source if the attP site stock does not contain it. If the screening marker is fluorescence-related, we may use the genomic P{nos-phiC31\int.NLS}X source (non-fluorescence), please let us know if this is the case.
For vermillion+ markers, e.g. for creating stable attP-gRNA lines, you may actually choose any of the vermillion+ based, non-X, attP lines from these below lists. We may setup the crosses to the y-v- line and screen for v+. You may get the same v+ G2 transformants as using the v- based attP lines. This is just regular Plan H or Plan I with extra v+ screening. You may also ask us to keep the resulting gRNA lines and have us cross to any of the Cas9 stocks and inject their F1s (additional Plan R) later.
P{CaryP}
Two attP landing sites were created by the Calos group. These two sites were initially created by the traditional P-element system. See below paper for more info.
PhiC31-mediated RMCE
In June, 2006, the Wu lab published a method that targets constructs to predetermined genomic sites using the phiC31 integrase system in conjunction with Recombinase Mediated Cassette Exchange (RMCE).
Please visit the Wu lab for more info:
http://transvection.org
P[acman]
P[acman] system makes use of the combined tools of a conditional amplifiable technique for typical P1 and BAC construct, recombineering, and PhiC31-mediated trangenesis. It has the advantage of delivering large DNA fragments into specific genomic locations with attP site, which were generated by the Ballen lab and markered by yellow. Here we are incorporating the P[acman] system with the work from the Basler group which used endogenous PhiC31 source instead of co-injecting the PhiC31 mRNA.
Please visit the Bellen lab for more info:
http://flypush.imgen.bcm.tmc.edu/lab/index.html
MiMIC
MiMIC is a highly versatile transposon insertion resource for engineering Drosophila melanogaster genes. MiMIC system was developed by the collaboration among the laboratories of Hugo Bellen (Baylor College of Medicine), Roger Hoskins (Lawrence Berkeley National Laboratory) and Allan Spradling (Carnegie Institution of Washington).
Please visit the Bellen lab MiMIC page for more info:
http://flypush.imgen.bcm.tmc.edu/pscreen/technique.html
FlyC31
In FlyC31 system, a library of landing platforms containing attP site throughout the Drosophila genome were created by the Basler group, which are ready for the transgenesis of the construct with attB site. These landing platforms were deliberately designed to be manipulated in vivo by Cre/loxP system after transgenesis to get rid of most of the non-necessary sequences and only keep your gene-of-interest flanking with one loxP and one attL site (We do not provide this Cre/loxP service).
Please visit the FlyC31 website for more info:
http://www.flyc31.org/why_phic31.php
Perrimon Strains
Additional attP landing sites were created by the Perrimon group, and were tested for position effects by luciferase expression on various tissues. See below paper for more info.
Please visit the Perrimon lab website for more info:
http://genetics.med.harvard.edu/~perrimon/
The PhiC31 system and the Gateway System
One of the "Gateway-PhiC31 system", for example, can be found here. Please note the attP1, attP2, attB1, attB2, etc, at the original Murphys Gateway are of different concepts.
Reference:
Groth A, Fish M, Nusse R, Calos MP (2004) Construction of transgenic Drosophila by using the site-specific integrase from phage PhiC31. Genetics 166: 1775
Bateman JR, Lee AM, Wu C-t (2006) Site-specific transformation of Drosophila via PhiC31 integrase-mediated cassette exchange. Genetics 173:769
Venken KJT, He Y, Hoskins RA, Bellen HJ (2006) PhiC31: a BAC transgenic platform for targeted insertion of large DNA fragments in Drosophila melanogaster. Science 314:1747
Bischof J, Maeda RK, Hediger M, Karch F, Basler K (2007) An optimized transgenesis system for Drosophila using germ-line-specific PhiC31 integrases. PNAS 104:3312
Markstein M, Pitsouli C, Villalta C, Celniker SE, Perrimon N (2008) Exploiting position effects and the gypsy retrovirus insulator to engineer precisely expressed transgenes. Nature Genetics 40:476
Knapp J-M, Chung P, Simpson JH (2015) Generating Customized Transgene Landing Sites and Multi-Transgene Arrays in Drosophila Using phiC31 Integrase. Genetics 199: 919
Currently available Calos strains (P{CaryP} strains) at BestGene Inc.
Site name |
BDSC Stock# |
Genotype |
Estimated CytoSite# |
Transformant Eye Color1 |
Transformation Score [Avg. (BACs)]6 |
attP1 |
8621 |
y1 w67c23; P{CaryP}attP1 |
55C4 |
Red |
6.4 (2.2) |
attP2 |
8622 |
y1 w67c23; P{CaryP}attP2 |
68A4 |
Red |
5.8 (1.9) |
attPY |
41124 |
y1 w*/Dp(2;Y)G, P{CaryP}attPY |
Y |
Red |
4.3 (nd) |
#FlyBase Drosophila melanogaster (R6.42)
1Eye color were determined three days after eclosion (heterozgous state)
6Transformation efficiency Score was calculated based on actual running orders and will be updated quarterly. Average score and >30kb BACs scores were shown
Currently available Wu strains and Bateman strains (PhiC31-RMCE strains) at BestGene Inc.
Site name |
BDSC Stock# |
Genotype |
Estimated CytoSite# |
Original Stock Eye Color7 |
Transformation Score6 |
JB37B |
27387 |
P{attP.w[+].attP}JB37B |
37B7 |
Red |
6.8 |
JB38F |
27388 |
P{attP.w[+].attP}JB38F |
38F1 |
Orange |
6.4 |
JB53F |
27386 |
P{attP.w[+].attP}JB53F |
53F8 |
Orange |
3.8 |
JB89B |
25091 |
P{attP.w[+].attP}tara[JB89B] |
89B8 |
Red |
5.8 |
38451 |
38451 |
w1118; Dr1/TM3, P{attP.w+.attP}FS10 Sb1 Ser1 |
79A2 |
Orange |
3.3 |
38452 |
38452 |
w1118; Dr1/TM3, Sb1 Ser1 P{attP.w+.attP}FS18 |
100D1 |
Orange |
2.0 |
38453 |
38453 |
y1 w*; snaSco/CyO, P{attP.w+.attP}J04 |
37B8 |
Orange |
9.0 |
38454 |
38454 |
y1 w*; snaSco/CyO, P{attP.w+.attP}J08 |
37F2 |
Red |
4.6 |
38455 |
38455 |
FM7h, P{attP.w+.attP}FS2/C(1)DX, y1 f1 |
10B6 |
Red |
4.6 |
38456 |
38456 |
FM7h, P{attP.w+.attP}FS5/C(1)DX, y1 f1 |
6F3 |
Orange |
3.0 |
#FlyBase Drosophila melanogaster (R6.42)
7Eye color were determined three days after eclosion. This is the color of the original stock, not the transformant
6Transformation efficiency Score was calculated based on actual running orders and will be updated quarterly. Average score and >30kb BACs scores were shown
Currently available Bellen strains (P[acman] strains) at BestGene Inc.
Site name |
BDSC Stock# |
Genotype |
Estimated CytoSite# |
Transformant Eye Color1 |
Transformation Score [Avg. (BACs)]6 |
VK10 |
97292 |
PBac{yellow[+]-attP-3B}VK00010 |
chromosome 2 |
- |
- (nd) |
VK15 |
9734 |
PBac{yellow[+]-attP-9A}VK00015 |
chromosome 2 |
Red |
3.3 (2.4) |
VK38 |
9753 |
PBac{yellow[+]-attP-3B}VK00038 |
5B8 |
Red |
4.1 (0.8) |
VK06 |
9726 |
PBac{yellow[+]-attP-9A}VK00006 |
19E7 |
Light Orange |
1.9 (1.3) |
VK37 |
9752 |
PBac{yellow[+]-attP-3B}VK00037 |
22A3 |
Orange |
5.8 (2.4) |
VK12 |
9731 |
PBac{yellow[+]-attP-9A}VK00012 |
25A3 |
Light Orange |
1.1 (nd) |
VK02 |
9723 |
PBac{yellow[+]-attP-3B}VK00002
| 28E7 |
Red |
6.4 (2.7) |
VK11 |
9730 |
PBac{yellow[+]-attP-3B}VK00011 |
40E4 |
Red |
0.4 (0.7) |
VK14 |
9733 |
PBac{yellow[+]-attP-9A}VK00014 |
43A1 |
Orange |
6.4 (2.7) |
VK16 |
9735 |
PBac{yellow[+]-attP-3B}VK00016 |
47C6 |
Orange |
5.0 (3.2) |
VK30 |
9747 |
PBac{yellow[+]-attP-9A}VK00030 |
50E1 |
Orange |
3.7 (1.4) |
VK18 |
9736 |
PBac{yellow[+]-attP-9A}VK00018 |
53B2 |
Orange (maybe variegated) |
6.6 (2.6) |
VK22 |
9740 |
PBac{yellow[+]-attP-9A}VK00022 |
57F5 |
Orange |
6.9 (2.9) |
VK01 |
9722 |
PBac{yellow[+]-attP-3B}VK00001 |
59D3 |
Light Orange |
4.8 (3.8) |
VK31 |
9748 |
PBac{yellow[+]-attP-3B}VK00031 |
62E1 |
Orange |
6.3 (2.8) |
VK33 |
9750 |
PBac{yellow[+]-attP-3B}VK00033 |
65B2 |
Red |
6.2 (3.5) |
VK39 |
9754 |
PBac{yellow[+]-attP-3B}VK00039 |
67E4 |
- |
4.8 (nd) |
VK19 |
9737 |
PBac{yellow[+]-attP-9A}VK00019 |
68D2 |
Red |
4.2 (1.9) |
VK23 |
9741 |
PBac{yellow[+]-attP-9A}VK00023 |
70A2 |
Red |
4.7 (3.8) |
VK36 |
9751 |
PBac{yellow[+]-attP-3B}VK00036 |
70C4 |
Red |
4.0 (nd) |
VK05 |
9725 |
PBac{yellow[+]-attP-9A}VK00005 |
75A10 |
Orange |
7.2 (2.2) |
VK13 |
9732 |
PBac{yellow[+]-attP-9A}VK00013 |
76A2 |
Red |
6.0 (4.4) |
VK21 |
9739 |
PBac{yellow[+]-attP-9A}VK00021 |
76C5 |
Orange |
- (nd) |
VK07 |
9727 |
PBac{yellow[+]-attP-3B}VK00007 |
82A1 |
Orange |
2.7 (0.0) |
VK32 |
9749 |
PBac{yellow[+]-attP-3B}VK00032 |
85A2 |
Light Orange |
2.4 (0.0) |
VK40 |
9755 |
PBac{yellow[+]-attP-3B}VK00040 |
87B10 |
Red |
7.2 (1.4) |
VK27 |
9744 |
PBac{yellow[+]-attP-9A}VK00027 |
89E11 |
Red |
7.6 (1.8) |
VK26 |
9743 |
PBac{yellow[+]-attP-9A}VK00026 |
96F3 |
Light Orange |
2.71 (3.8) |
VK20 |
9738 |
PBac{yellow[+]-attP-9A}VK00020 |
99F8 |
Light Orange |
4.8 (3.6) |
VK24 |
97422 |
PBac{yellow[+]-attP-9A}VK00024 |
101F1 |
- |
0.0 (nd) |
#FlyBase Drosophila melanogaster (R6.42)
1Eye color were determined three days after eclosion (heterozgous state)
2These lines were very hard to transform
6Transformation efficiency Score was calculated based on actual running orders and will be updated quarterly. Average score and >30kb BACs scores were shown
Currently available Basler strains (FlyC31 strains) at BestGene Inc.
Site name |
Reference BDSC Stock# |
Genotype |
Estimated CytoSite# |
Transformant Eye Color3 |
Transformation Score [Avg. (BACs)]6 |
ZH-2A |
24480 |
M{3xP3-RFP.attP'}ZH-2A |
2A3 |
Orange |
3.3 (0.6) |
ZH-22A |
24481 |
M{3xP3-RFP.attP'}ZH-22A |
22A2 |
Light Orange |
5.2 (3.4) |
ZH-51C |
24482 |
M{3xP3-RFP.attP'}ZH-51C |
51C1 |
Light Orange |
7.5 (4.9) |
ZH-51D |
244835 |
M{3xP3-RFP.attP}ZH-51D |
51D9 |
Red |
6.2 (2.4) |
ZH-58A |
244844 |
M{3xP3-RFP.attP}ZH-58A |
58A3 |
Light Orange |
7.0 (5.5) |
ZH-68E |
24485 |
M{3xP3-RFP.attP'}ZH-68E |
68E1 |
Light Orange |
6.8 (2.9) |
ZH-86Fa |
244864 |
M{3xP3-RFP.attP'}ZH-86Fa |
86E18 |
Red |
7.0 (3.8) |
ZH-86Fb |
24749 |
M{3xP3-RFP.attP}ZH-86Fb |
86F8 |
Red |
7.3 (4.3) |
ZH-102D |
2448813 |
M{3xP3-RFP.attP}ZH-102D (with M{vas-int.Dm}ZH-2A) |
102F4 |
Red |
7.3 (3.4) |
24480NF |
- |
24480NF12 ZH-2A |
2A3 |
Orange |
2.0 (nd) |
24481NF |
- |
24481NF11 ZH-22A |
22A2 |
Light Orange |
- (0.0) |
24482NF |
- |
24482NF11 ZH-51C |
51C1 |
Orange |
2.0 (nd) |
24483NF |
- |
24483NF11 ZH-51D |
51D9 |
Red |
3.4 (nd) |
24484NF |
- |
24484NF11 ZH-58A |
58A3 |
Light Orange |
1.3 (nd) |
24485NF |
- |
24485NF11 ZH-68E |
68E1 |
Light Orange |
1.1 (0.0) |
24486NF |
- |
24486NF11 ZH-86Fa |
86E18 |
Red |
5.9 (nd) |
24749NF |
- |
24749NF11 ZH-86Fb |
86F8 |
Red |
8.3 (2.4) |
|
|
|
#FlyBase Drosophila melanogaster (R6.42)
3According to FlyC31 website
4These strains are weak; BDSC#24486 is the weakest strain (low survival rate after injection)
5Enhancer trap effect from a nearby gene hibris and is not useful for reporter gene analysis and other applications where ectopic expression is an issue
6Transformation efficiency Score was calculated based on actual running orders and will be updated quarterly. Average score and >30kb BACs scores were shown
11M{vas-int.Dm}ZH-2A removed and loxP-3xP3-RFP removed version
12M{vas-int.Dm}ZH-102D removed and loxP-3xP3-RFP removed version
13Plan I is NOT available for this stock
Currently available Perrimon strains (P{CaryP} strains) at BestGene Inc.
Site name |
BDSC Stock# |
Genotype |
Estimated CytoSite# |
Transformant Eye Color1 |
Transformation Score [Avg. (BACs)]6 |
attP18 |
32107 |
y1 w67c23 P{CaryP}attP18 |
6C12 |
Orange |
4.0 (0.0) |
attP4 |
- |
y1 w67c23 P{CaryP}attP4 |
12C6 |
Orange |
2.5 (0.0) |
attP3 |
32230 |
y1 w67c23 P{CaryP}attP3 |
19C4 |
Light Orange |
0.9 (0.7) |
attP40 |
- |
y1 w67c23; P{CaryP}attP40 |
25C6 |
Red |
7.2 (3.7) |
attP30 |
- |
y1 w67c23; P{CaryP}attP30 |
29C3 |
Red |
4.0 (nd) |
attP14 |
- |
y1 w67c23; P{CaryP}attP14 |
36A10 |
Orange |
4.0 (1.0) |
attP88 |
- |
y1 w67c23; P{CaryP}attP88 |
64A12 |
Red |
4.4 (nd) |
attP112 |
- |
y1 w67c23; P{CaryP}attP112 |
68C13 |
Red |
3.0 (nd) |
attP64 |
- |
y1 w67c23; P{CaryP}attP64 |
89B9 |
Red |
5.3 (nd) |
attP154 |
- |
y1 w67c23; P{CaryP}attP154 |
97D2 |
Orange |
6.3 (1.8) |
#FlyBase Drosophila melanogaster (R6.42)
1Eye color were determined three days after eclosion (heterozgous state)
6Transformation efficiency Score was calculated based on actual running orders and will be updated quarterly. Average score and >30kb BACs scores were shown
Other strains*
Site name |
BDSC Stock# |
Genotype |
Estimated CytoSite# |
Transformant Eye Color1 |
Transformation Score [Avg. (BACs)]6 |
su(Hw)attP8 |
32233 |
y1 w* P{CaryIP}su(Hw)attP8 |
8E10 |
Red |
3.8 (0.0) |
su(Hw)attP6 |
347679 |
y1 w* P{CaryIP}su(Hw)attP6 |
24A2 |
Red |
5.0 (nd) |
su(Hw)attP5 |
34765 |
y1 w* P{CaryIP}su(Hw)attP5 |
50F1 |
Red |
6.1 (3.8) |
su(Hw)attP4 |
34763 |
y1 w* P{CaryIP}su(Hw)attP4 |
67E2 |
Red |
3.0 (nd) |
su(Hw)attP2 |
347619 |
y1 w* P{CaryIP}su(Hw)attP2 |
92D9 |
Red |
4.8 (nd) |
VIE260B |
VDRC#60100 |
P{attP,y+,w3'}VIE-260B |
40D3 |
Red |
8.5 (nd) |
JK22C aka JMK-96 |
- |
w1118; P{CaryP}JK22C |
22C3 |
Red |
5.9 (0.0) |
JK65C aka JMK-98 |
- |
w1118; P{CaryP}JK65C |
65C3 |
Red |
5.6 (0.0) |
JK66B aka JMK-99 |
- |
w1118; P{CaryP}JK66B |
66B4 |
Red |
3.8 (1.2) |
*Some lines contain the nos-PhiC31 genomic integrase source as indicated at the Genotype.
#FlyBase Drosophila melanogaster (R6.42)
1Eye color were determined three days after eclosion (heterozgous state)
6Transformation efficiency Score was calculated based on actual running orders and will be updated quarterly. Average score and >30kb BACs scores were shown
9This strain has very high non-specific integration rate. Service W (PCR confirmation of integration site) is highly recommended
updated 9/27/2024
Highlighted are popular/preferred strains
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