# Analysis steps on tutorial

**URL:** <https://forum.mothur.org/t/analysis-steps-on-tutorial/21280>\
**Category:** Commands in mothur\
**Created:** [February 19, 2022, 12:09pm UTC](https://forum.mothur.org/t/analysis-steps-on-tutorial/21280 "2022-02-19T12:09:58Z")\
**Posts on this page:** 3\
**Page:** 1

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**Author:** ![junkim83](https://avatars.discourse-cdn.com/v4/letter/j/bcef8e/32.png) [@junkim83](https://forum.mothur.org/u/junkim83)\
**Post date:** [February 19, 2022, 12:09pm UTC](https://forum.mothur.org/t/analysis-steps-on-tutorial/21280/1 "2022-02-19T12:09:58Z")

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Hi,

Sorry for too many post.  
Since I checked tutorial again. I found something wired.

On analysis steps, based on tutorial…

```auto
mothur > count.groups(shared=final.opti_mcc.shared)
mothur > sub.sample(shared=final.opti_mcc.shared, size=2403)
mothur > rarefaction.single(shared=final.opti_mcc.shared, calc=sobs, freq=100)

```

However, when I ran sub.sample commands, it gave me the file named “final.opti\_mcc.0.03.subsample.shared”

Should rarefaction.single commans like below?

```auto
mothur > rarefaction.single(shared=final.opti_mcc.0.03.subsample.shared, calc=sobs, freq=100)

```

Best,  
Jun

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<div class="post-metadata">

**Author:** ![pschloss](https://yyz2.discourse-cdn.com/flex036/user_avatar/forum.mothur.org/pschloss/32/4_2.png) [@pschloss](https://forum.mothur.org/u/pschloss)\
**Post date:** [February 22, 2022, 5:41pm UTC](https://forum.mothur.org/t/analysis-steps-on-tutorial/21280/2 "2022-02-22T17:41:20Z")

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Nope. `rarefaction.single` needs to have all the data. It effecitvely runs sub.sample 1000 times and each time it calcualtes the alpha diversity metrics and at the end reports the average and standard deviation.

Pat

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<div class="post-metadata">

**Author:** ![system](https://yyz2.discourse-cdn.com/flex036/user_avatar/forum.mothur.org/system/32/2_2.png) [@system](https://forum.mothur.org/u/system)\
**Post date:** [March 4, 2022, 5:42pm UTC](https://forum.mothur.org/t/analysis-steps-on-tutorial/21280/3 "2022-03-04T17:42:12Z")

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