Frequently Asked Questions
Yes, we provide injections using customer’s flies. We charge an extra $25 for the special handling. Please send flies equivalent to two to three bottles of newly emerged flies. Please transfer these flies into fresh vials before sending to us.
There are w1118 and yw. You can choose one of them for your application.
We have integrase-combined lines and attp lines without integrase. It is better to choose integrase-combined lines because they have higher efficiency.
The integrase is on the X chromosome and is marked by GFP/RFP. The male transformants from the injected male fly cross should be integrase-free.
Yes, we will co-inject helper integrase DNA which we provide.
These attp lines work very well for the third chromosome: 86Fb, 86Fa, 24871 (or 9750), 35569 (or 9744), R8622 (or 25710 in vermilion background).
These attp lines work very well for the second chromosome: 51C, 51D, 24872 (or 9752), attp40w (25709 vermilion background), 9736
There are not many attp lines available for X chromosome and the efficiency is not very good. You can try one of these lines: 32107, 32230, 32233, ZH-2A
You can use TE or EB buffer for P-element- or piggyBac-based injections. For phiC31-based injections (including MiMIC and RMCE), please use DEPC water or EB buffer so we can directly dilute the sample into the right concentration and inject.
You need to provide a MiMIC line and plasmid. We will co-inject your plasmid with helper integrase to the MiMIC line you provide. It may take a couple of weeks to expand the flies. The turn-around time depends on how many flies you send to us.
We will use yw double balancer to set up the cross and screen for loss of y+ flies
We have many Cas9-expressing lines, such as nosCas9 attp2, nosCas9 attp40, 51324, 54591, etc. These lines all work well and you can decide which line you want to use.
Either way works. If you premix your sample, we need at least 20ul total volume per injection. If you send samples individually, we need 20ug to 30ug total DNA for each sample. The concentration for gRNA is over 0.5ug/ul and over 1.0ug/ul for donor.
Please use DEPC water or EB buffer.
Please use FedEx priority overnight mail to ship RNA, Oligos, and protein on dry ice.
Yes, we can co-inject with helper Cas9 plasmid, but it is not very efficient. We can also do the injections with Cas9 mRNA and Cas9 protein, which you need to provide.
Cas9 mRNA and Cas9 protein work very efficiently; however, Cas9 protein can cause some problems for injections since it is easy to clog the needles.
You can premix your sample with Cas9 protein at 300ng/ul to 500ng/ul, gRNA at 100ng/ul, donor at 250ng/l to 500ng/ul. We need total 20ul for each injection. Please send your sample on dry ice. In this way, you can save your sample.
Yes, we provide one free re-injection if you don’t get any transformants. However, for the injection of CRISPR/Cas9 or using customer’s flies, we only provide free re-injection if the survival rate is too low.