Method for evaluating diversity of marine zooplankton on basis of macro bar code technology
A zooplankton and diversity technology, applied in the direction of biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problems that the identification method is difficult to meet the operational monitoring, the professional skills of the identification personnel are high, and the species diversity is underestimated. , to achieve the effect of eliminating product purification steps, improving sequencing quality, and increasing resolution
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Embodiment 1
[0059] 1. Sample collection
[0060] In August 2017, 22 zooplankton stations ( figure 1 ) for sampling. The shallow water type II plankton net (mesh aperture 0.160mm, net opening diameter 31.6cm, net length 140cm) was used to collect zooplankton samples according to the "Marine Monitoring Specification" (GB 17378.7-2007). The samples collected at each station were thoroughly mixed and divided into two parts: one part was fixed with 5% formaldehyde for morphological identification, and the other part was fixed with absolute ethanol after filtering out seawater with a sieve of the same aperture. molecular identification.
[0061] 2. DNA Extraction of Sample Metagenome
[0062] Draw zooplankton samples three times from each sample preserved in alcohol (total volume 100mL), 10mL each time, combine the zooplankton samples drawn three times, and filter with a 20μm sieve to remove alcohol and residual planktonic algae Wait. DNA blood and tissue extraction kit (Qiagen Canada lnc....
Embodiment 2
[0072] The primers used for PCR amplification contain an 8-base tag sequence, which is a specific sequence. 22 primer pairs (see Table 2) are provided for PCR amplification of 22 samples, and the rest are completely consistent with Example 1.
[0073] Table 2 Primer pairs used in sample PCR amplification
[0074]
[0075]
Embodiment 3
[0077] The PCR reaction system is as follows (25μL): 1x Ex Taq Buffer (20mM Mg 2+ plus, Takara, Dalian, China), 5.0mM dNTP, 0.27mM trimethylamine oxide, 0.42μM 1,6-bis(N1-p-chlorophenyl-N5-biguanide) hexane digluconate, 15pmol containing Index Primers, 100ng of genomic DNA, and PCR products were not purified, and the rest were completely consistent with Example 2.
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