Lyles's blistering 9.86sec was the best time this year and a personal best, and lays down a marker ahead of the world championships in the autumn and the Tokyo Olympics next summer.
Noah Lyles beat fellow American Christian Coleman by just 0.006secs to win the 100 metres.
The South African, 28, who won gold in Rio in 2016 and London four years earlier will not lineup for the 800m after winning her most recent appearance over the distance in Doha on May 3.
Semenya's case has provoked a furious debate across sport worldwide about gender and "hyperandrogenic" athletes, those with "differences of sexual development" (DSD).
Semenya's case has provoked a furious debate across sport worldwide about gender and "hyperandrogenic" athletes.
Jamaica's male sprinters struggled at last year's Commonwealth Games, where Yohan Blake trailed in third in the 100 metres and the men's 4x100m relay team settled for bronze behind England and South Africa.
Geoffrey Kirui instead heads up the Kenyan men's quintet announced Tuesday while two-time title holder Edna Kiplagat leads the women's quartet for the Doha worlds, scheduled for September 27-October 6.
Kipchoge went close to smashing the barrier when he came in 25 seconds off the mark in a staged race around Italy's Monza race circuit in 2017.
This week South Africa middle-distance star Semenya lost a challenge against the International Association of Athletic Federations over plans to force some women to regulate their testosterone levels.
The Diamond League was held for the first time at the Khalifa International Stadium, which will host this year’s World Championships.
The Young Qatari’s time of 1min44.82sec at Doha Diamond League meeting easily surpassed the qualifying mark.
In the men’s 800m Qatari hope Addalla Abubaker Haydar put in an impressive performance but had to settle for a fourth place finish.
The South African stormed to victory in her final race before the CAS ruling on hyperandrogenism comes into effect on May 8.
IAAF's controversial rule regulating testosterone levels for women athletes will come into effect on May 8.
A cast including six reigning world champions, eight reigning Olympic champions and winners of nine 2018 Diamond Trophies will take to Khalifa International Stadium on Friday.