title: GSM8660179: SZ3; human saliva metagenome; OTHER
library_construction_protocol: 1. Add 0.8 times the volume of Vazyme VAHTSTM DNA Clean Beads into 25 μl PCR products, and after full suspension by shaking, adsorb them on the magnetic rack for 5 min, and carefully use the pipette to absorb the supernatant;  2. Add 20μl 0.8 times magnetic bead washing liquid, suspend fully after shaking, place on the magnetic rack for 5 min, and carefully suck out the supernatant;  3. Add 200μl 80% ethanol, place it on the magnetic rack in reverse, and adsorb it to the other side of the PCR tube with magnetic beads, and absorb the supernatant after full adsorption;  4, put it at room temperature for 5 minutes, until the alcohol volatilization is complete, the magnetic beads can be cracked;  5. Add 25μl Elution Buffer for elution;  6. Put the PCR tube on the adsorption rack for 5 min, fully adsorb, and remove the supernatant into a clean 1.5ml centrifuge tube for storage. llumina's TruSeq Nano DNA LT Library Prep Kit was used to prepare sequencing libraries.  1) The amplified products were repaired at the End of the sequence first. The protruding base at the 5 'end of the DNA sequence was removed through End Repair Mix 2 in the kit, and a phosphate group was added to supplement the missing base at the 3' end.  2) Add A base to the 3 'end of the DNA sequence to prevent the DNA segment from self-connecting, while ensuring that the target sequence can be connected to the sequencing connector (the 3' end of the sequencing connector has a prominent T base);  3) A sequencing connector containing a library specific tag (i.e., Index sequence) is added to the 5 'end of the sequence, so that DNA molecules can be fixed on the Flow Cell;  4) BECKMAN AMPure XP Beads were used to remove the joint self-connecting segments through magnetic bead screening and purify the library system after adding the joint;  5) PCR was performed on the DNA fragments connected with the above connectors to enrich the sequencing library template, and BECKMAN AMPure XP Beads were used to purify the enriched products of the library again.  6) The final fragment selection and purification of the library was performed by 2% agarose gel electrophoresis.