Decide to try source, DNA removal, and you may genome sequencing

Decide to try source, DNA removal, and you may genome sequencing

Here by the entire genome sequencing out of 55 honey bees and by building a premier solution recombination chart during the honey bee, we learned that crossovers was for the GC stuff, nucleotide diversity, and gene density. I also affirmed the former suggestion you to family genes conveyed when you look at the staff member brains enjoys strangely highest CO pricing. Our studies hold the see you to definitely variation out of staff member decisions, although not protected setting, was a motorist of the highest crossing-more price when you look at the bees. We discover zero proof your crossing-more than speed is actually with a premier NCO price.

Methods and information

Four territories off honeybees (Apis mellifera ligustica Spin) was in fact obtained regarding a great bee ranch within the Zhenjiang, Asia. For each nest contained you to queen, those drones, and you can hundreds of workers. Bees out of about three colonies was in fact picked to own entire genome sequencing.


Brand new DNA of each and every individual is extracted having fun with phenol/chloroform/isoamyl alcoholic beverages strategy. To minimize the possibility of bacterial toxic contamination, the newest abdomens regarding bees was basically eliminated ahead of DNA removal. Throughout the step 3 ?g from DNA away from for every shot were utilized to have whole genome resequencing since left DNA are leftover for PCR and Sanger sequencing. Build of DNA libraries and you will Illumina sequencing was indeed did at BGI-Shenzhen. Inside the short-term, paired-prevent sequencing libraries which have enter size of five-hundred bp was indeed created for each and every take to according to manufacturer’s directions. Up coming 2 ? a hundred bp paired-end checks out was made on the IlluminaHiSEq 2000. The new queens had been sequenced at the everything 67? publicity on average, drones within up to 35? publicity, and you can pros at the around 30? exposure (Desk S1 into the More file dos). The fresh sequences was in fact deposited regarding the GenBank database (accession no. SRP043350).

SNP contacting and you can marker personality

Honeybee source genome is actually downloaded regarding NCBI . The sequencing reads had been very first mapped onto source genome with bwa then realigned which have stampy . Up coming regional realignment up to indels try did by Genome Research Toolkit (GATK) , and alternatives had been named because of the GATK UnifiedGenotyper.

Considering the down reliability away from contacting indel alternatives, simply known SNPs are utilized due to the fact indicators. Very first, 920,528 to 960,246 hetSNPs have been entitled in the for every queen (Desk S2 for the Even more file dos). After that, around twenty-two% ones was removed because the internet sites also are hetSNPs within the one or more haploid drone (this might reflect non-allelic succession alignments as a result of CNVs, sequencing mistake, otherwise lower sequencing quality). Equivalent dimensions of this new hetSNPs along with had been seen in human jizz sequencing . In the end, 671,690 in order to 740,763 reputable hetSNPs during the per colony were used just like the indicators to place recombination incidents (Desk S2 into the Extra file 2).

Haploid phasing

For each colony, the identified markers were used for haploid phasing. The linkage of every two adjacent markers was inferred to determine the two chromosome haplotypes of the queen by comparing the SNP linkage information across all drones from the same colony. Detailed methods were described in Lu’s study . In brief, for each pair of adjacent hetSNPs, for example A/G and C/T, there could be two types of link in the queen ‘A-C, G-T’ or ‘A-T, G-C’. Assuming recombination events are low probability, if more ‘A-C, G-T’ drones are found than ‘A-T, G-C’ drones, then ‘A-C, G-T’ is assumed to be the correct link in the queen and vice versa. The two haplotypes can be clearly discriminated between >99% of ple). For linkage of the <1% markers, as shown in Additional file 1: Figure S2B, between markers at ‘LG1:20555174' and ‘LG1:20555456' , there are 14 ‘A-A or G-G' type drones against 1 ‘A-G or G-A' type drone, so ‘A-A, G-G' is assumed to be the correct link in queen and a recombination event is identified at this site in sample I-9.

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