Fair enough. I guess I was overreacting a bit to the "bullet proof skin" BS. Some hard data is what we need. That's what I'm hoping for from the results of the two rounds of genetic modifications.
An important point about the two step process: zinc fingers generally have a monoallelic effect, not biallelic. That means that usually only one of the two copies gets the sequence change. So if the gene for worm silk protein (WSP) was removed, the resulting worm would still have the other copy present and would produce WSP, just half as much of it. The worms would then need to be crossed to get worms homozygous for the WSP deletetion before they would make pure spider silk with no WSP at all. The silk with 1/2 level of WSP would be superior to Monster Silk and much closer to pure spider silk but not pure spider silk. (I would calculate about 90% of the strength of pure spider silk which is just very close. That's estimated assuming the effect is linear. BUT if SSP sticks to itself better than it does to WSP (quite possible) then the silk will be stronger, maybe as much as 95%. (Note that it's quite possible that the Monster Silk worms may have 2, 3 or even more SSP genes added. (I would say AT LEAST TWO which would be why Monster Silk was superior to the other silks in those 20 lines.) If you had 3 SSP genes added you'd then have 6 copies producing SSP and only one copy producing WSP (the other having been knocked out) Recall that worm silk is only 40% of the strength of pure spider silk. But after crossing to make homozygous it should be 100%.)
[[Note: this is a very general tendency in agricultural animals that have been bred for maximum expression of particular traits. Often the traits are expressed so strongly because the animals have multiple copies of the gene that produces the protein that controls the trait.]]
That may be why Kim said "expect results in the early fall" Silk with only 1/2 the usual amount of WSP would definitely require testing of the protein levels to confirm.
Note that knocking out one of the two copies of the WSP gene would be 99.999% of the way to success. Knocking out one copy is the hard part. Once you have worms with only one copy you just cross them and 1/4 of the offspring have NO copies. Dead easy, dead routine, only a matter of time (one generation).
So what could be going on is 1st round knock out the WSP gene, cross them to get worms with NO WSP gene in either copy at the same time you do the 2nd round of additional GMs to either the eggs of that cross or to the ovaries of the female moths before crossing them. So the results of the second round could be (1/4 of them) homozgyous for the WSP deletion (NO WSP) AND heterozygous for the additional GMs.
NOTE if the GMs are done by injection into the ovaries ALL of the worms getting the GMs will necessarily be homozygous because the copy of the gene coming from the father will be unaffectd. OTOH if the injections are done to single cell eggs that have already been fertilized then a small percentage would be already homozygous for the GM. (Not all worms take up the GM. that's why you get 10 aliquots of zinc fingers in a set. With 10 tries you should get about 3 to 5 with that take up the GM in one copy of the gene. A smaller percentage will take it up in both copies, so IF you are a bit lucky you'll get a worm with the GM in both copies on the first round. If you get REALLY lucky you might get both a male and a female with the GM in both copies (one can dream!) Then you could start ramping up the population right away.
So what we might see is pure spider silk produced by worms that were crosses of the first round of GMs PLUS the additional GMs: worms producing pure spider silk plus { worms producing pure spider silk with a hepcidin antibotic add-on as well, or worms producing the artificial sequence; EITHER of which would constitute proof of principle of the "platform worm".
In the 8 August PR Kim said "The current second round is focused on two objectives: 1) modifying the profile of the expressed proteins in the previously disclosed "Monster Silk" fiber, and 2) potentially creating a new recombinant silk platform technology."
" modifying the profile of the expressed proteins" in Monster silk could mean that it was in THIS round that they knocked out the WSP gene, which, as I noted, would not produce pure spider silk immediately unless they got lucky and got a worm with both copies knocked out. Otherwise they'd get a worm producing 1/2 the previous level of WSP (which is a modified profile of the expressed protein). If that's the case I'm not sure what they'd have done in the first round - Maybe the artificial sequence, which could then be made homozygous by the time the second round was complete.
OTOH " modifying the profile of the expressed proteins " could mean a LOT of other things which I suspect is exactly why KBLB chose that wording. They are keeping exactly what they are doing VERY close to the vest here.
Note that all this is still considerably simplified. That's one on the reasons why it's wise to avoid getting very specific expectations about the pace and timing of developments. There is almost always a lot more going on than we are aware of so the timing may not be what we expect it to be. Patience is required.
AI
