2.1 Sample collection
S. chinensis samples were
collected in October, 2019, from fresh mature galls on R.
chinensis , in Wufeng county (30°10′ N,110°52′ E,960 m above sea
level), Hubei Province, China. S. chinensis individual for
sequencing were from a single gall for PacBio sequencing, Illumina
sequencing, RNA-seq and Hi-C analysis. All aphids within the gall were
presumed to be clonal offspring of a single fundatrix, because allS. chinensis galls have contained only a single fundatrix and the
fundatrix produces offspring in the gall via parthenogenesis.
Fundatrigeniae (female) contained within a gall were moved to a petri
dish after dissecting the gall. Impurities such as waxes were removed
manually. Samples were separated into randomly groups with each group
containing about 20 individuals. The samples were immediately frozen in
liquid nitrogen for 2 hours and then stored at -80℃ f for later
analysis.
In addition to the original samples, a colony was established through
artificial cultivation for other genetic studies. To establish a colony,
autumn migrants of S. chinensis were collected from mature galls
and transferred to a nursery of the moss Plagiomnium
maximoviczii , which were maintained in a greenhouse. In the following
year, spring migrants (sexuparae) were collected from the mosses and
cultivated in lab. Aphids were transferred to host trees after fundatrix
emergence for gall induction. Aphid samples of different stages
including fundatrix, fundatrigeniae, autumn migrants, overwinter nymphs,
and spring migrants as well as male and female sexuales were collected
separately. All aphid samples were immediately frozen in liquid nitrogen
for two hours and then stored at -80℃ until further analysis.