Improved production of recombinant aav using embryonated avian eggs

The method of any one of the preceding claims, wherein the AAV is of mammalian origin.

The method of any one of the preceding claims, wherein the first and second nucleic acid vectors are combined with a cationic polymer prior to inoculation.

The method of any one of the preceding claims, wherein the egg is incubated for a period of at least 15 hours, at least 20 hours, at least 24 hours, at least 36 hours, at least 54 hours, at least 60 hours, at least 72 hours, at least 80 hours, or at least 90 hours.

The method of any one of the preceding claims, wherein the rAAV is isolated using a manual pipette or syringe or a machine-controlled pipette or syringe.

The method of any one of the preceding claims, wherein the AAV is of an AAV1, an AAV2, an AAV3, an AAV4, an AAV5, an AAV6, an AAV7, an AAV8, an AAV9, an AAVrh10, an AAV12, or an AAV13 serotype.

The method of any one of the preceding claims, wherein the AAV is of an AAV1-M3, an AAV2-M3, an AAV2(tripYF), an AAV2(quadYF), an AAV2(pentaYF), an AAV2-BCDG(T491V+K556R), an AAV5-M2, an AAV5(Y719F), an AAV6(T492V+S663V), an AAV6(T492V+Y705F+Y731F), an AAV6(S551V+S663V), an AAV8-C&G(T494V), an AAV8-M3, an AAV8(Y733F), an AAV8(T494V+Y733F), AAV8(Y275F+Y447F+Y733F) or an AAV9-PHP.B serotype;

• or the serotype of another capsid variant.

The method of any one of the preceding claims, wherein an allantoic cavity, a chorioallantoic membrane, a yolk sac, or an amnion of the egg is inoculated.

The method of any one of the preceding claims further comprising:

• iv) producing purified rAAV by subjecting the isolated rAAV to an iodixanol gradient.

The method of any one of the preceding claims, wherein the egg is incubated in a shaker, a spinner or an automatic eggs incubator.

The method of any one of the preceding claims, wherein the transgene encodes a therapeutic protein.

An embryonated avian egg comprising a recombinant AAV of mammalian origin, wherein the recombinant AAV comprises a transgene.