BestGene Inc. - PhiC31 Info

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PhiC31 integrase-mediated transgenesis systems - P[acman] and FlyC31

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).

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

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.

Please visit the FlyC31 website for more info:
http://www.frontiers-in-genetics.org/flyc31/

Reference:

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

Currently available P[acman] strains at BestGene Inc.

BSC Stock#	Genotype	Estimated CytoSite	Coordinate	Eye Color¹
9729²	PBac{yellow[+]-attP-3B}VK00010	chromosome 2		-
9734	PBac{yellow[+]-attP-9A}VK00015	chromosome 2		Red
9753	PBac{yellow[+]-attP-3B}VK00038	5B8	X:5497935	Red
9726	PBac{yellow[+]-attP-9A}VK00006	19E7	X:20689962	Light Orange
9752	PBac{yellow[+]-attP-3B}VK00037	22A3	2L:1584486	Orange
9731	PBac{yellow[+]-attP-9A}VK00012	25A3	2L:4683203	Light Orange
9723	PBac{yellow[+]-attP-3B}VK00002	28E7	2L:8133094	Red
9724	PBac{yellow[+]-attP-3B}VK00003a	31B1	2L:10220182	Red
9724	PBac{yellow[+]-attP-3B}VK00003b	40A4	2L:21639162	Orange
9730	PBac{yellow[+]-attP-3B}VK00011	40E4	2L:21951742	Red
9733	PBac{yellow[+]-attP-9A}VK00014	43A2	2R:2285107	Orange
9735	PBac{yellow[+]-attP-3B}VK00016	47C4	2R:596343	Orange
9728	PBac{yellow[+]-attP-3B}VK00008	50A3	2R:8443624	Light Orange
9747	PBac{yellow[+]-attP-9A}VK00030	50E1	2R:9203330	Orange
9736	PBac{yellow[+]-attP-9A}VK00018	53B1	2R:11361979	Red
9740	PBac{yellow[+]-attP-9A}VK00022	57F5	2R:16689522	Orange
9722	PBac{yellow[+]-attP-3B}VK00001	59D3	2R:18279750	Orange
9746²	PBac{yellow[+]-attP-9A}VK00029	h47-h53	94181 of AABU01002142	-
9748	PBac{yellow[+]-attP-3B}VK00031	62E1	3L:2376116	Orange
9750	PBac{yellow[+]-attP-3B}VK00033	65B2	3L:6402205	Red
9754²	PBac{yellow[+]-attP-3B}VK00039	67E4	3L:10463229	-
9737	PBac{yellow[+]-attP-9A}VK00019	68D2	3L:11654656	Red
9741	PBac{yellow[+]-attP-9A}VK00023	70A2	3L:13170566	Red
9751	PBac{yellow[+]-attP-3B}VK00036	70C4	3L:13802829	Red
9725	PBac{yellow[+]-attP-9A}VK00005	75B1	3L:17900987	Orange
9732	PBac{yellow[+]-attP-9A}VK00013	76A2	3L:19153237	Red
9739	PBac{yellow[+]-attP-9A}VK00021	76C5	3L:19713047	Orange
9727	PBac{yellow[+]-attP-3B}VK00007	82A1	3R:81372	Orange
9749	PBac{yellow[+]-attP-3B}VK00032	85A2	3R:4301078	Light Orange
9755	PBac{yellow[+]-attP-3B}VK00040	87B10	3R:8265934	Red
9744	PBac{yellow[+]-attP-9A}VK00027	89E11	3R:12878600	Red
9745	PBac{yellow[+]-attP-9A}VK00028	92F1	3R:16375389	Orange
9743	PBac{yellow[+]-attP-9A}VK00026	96F3	3R:21662155	Light Orange
9738	PBac{yellow[+]-attP-9A}VK00020	99F8	3R:26368401	Orange
9742²	PBac{yellow[+]-attP-9A}VK00024	102A3	4:79131	-

¹Eye colors were determined three days after eclosion (heterozgous state)
²These lines were very hard to transform

Currently available FlyC31 strains at BestGene Inc.

BSC Stock#	Genotype	Estimated CytoSite	Eye Color³
23648	M{3xP3-RFP.attP}ZH-86Fb (with M{vas-int.B}ZH-102D)	86F	Orange
23647	M{3xP3-RFP.attP}ZH-102D (with M{vas-int.B}ZH-86Fb)	102D	Red
24480	M{3xP3-RFP.attP'}ZH-2A (with M{vas-int.Dm}ZH-102D)	2A	Orange
24482	M{3xP3-RFP.attP'}ZH-51C (with M{vas-int.Dm}ZH-2A)	51C	Light Orange
24483	M{3xP3-RFP.attP}ZH-51D (with M{vas-int.Dm}ZH-2A)	51D	Red
24484	M{3xP3-RFP.attP}ZH-58A (with M{vas-int.Dm}ZH-2A)	58A	Light Orange
24485	M{3xP3-RFP.attP'}ZH-68E (with M{vas-int.Dm}ZH-2A)	68E	Light Orange
24486⁴	M{3xP3-RFP.attP'}ZH-86Fa (with M{vas-int.Dm}ZH-2A)	86F	Red
24487	M{3xP3-RFP.attP'}ZH-96E (with M{vas-int.Dm}ZH-2A)	96E	Orange
24488	M{3xP3-RFP.attP}ZH-102D (with M{vas-int.Dm}ZH-2A)	102D	Red

³according to FlyC31 website
⁴BSC#24486 is the weakest strain (low survival rate after injection)